Symposium SM01—Materials for Biological and Medical Applications

Transmission electron microscopy (TEM) is the primary method to image biological cellular ultrastructure, though it is not possible with conventional TEM to label and distinguish different kinds of molecules in a single image. This serious limitation was recently addressed through the development of “multi-color EM,” which uses selective lanthanide ion tagging and electron energy-loss filtered imaging (Ramachandra et al., 2014, Microsc Microanal. 20; Adams et al., 2016, Cell Chem Biol. 23) yielding data analogous to multi-color fluorescence microscopy but at ~100x the magnification. While this technique promises to reveal novel structural information, it is tedious (requiring multiple long exposures with different energy filter settings), terribly inefficient (depending on the small fraction (<1%) of primary TEM electrons that are inelastically scattered), and yields noisy images with significantly lower resolution than is achievable by TEM imaging.

To improve the throughput, efficiency, and resolution of multi-color EM, we have developed a new multi-color EM technique with 4D-STEM, which uses a high-speed pixelated detector to record the vast majority of the primary electrons that interact with the specimen.

In order to achieve the necessary sensitivity for this technique, the pixelated detector must deliver synchronized (global shutter) readout of a large number of pixels (at least 512 × 512 pixels) and it must have single-electron sensitivity for electron counting.

We used the DE-16 direct detection camera in conjunction with the DE-FreeScan scan generator (Direct Electon LP, San Diego, CA USA) to collect a 4D-STEM dataset of a cellular mitomatrix sample, with mitochondria labeled by cerium and containing gold nanoparticles. After correcting for distortions in the diffraction patterns, we were able to develop a metric to distinguish the cerium labels and gold nanoparticles, while simultaneously generating bright-field and dark-field images of the specimen at significantly higher resolution than is possible through fluorescence light microscopy.

Neural electrodes have been widely used to monitor neural signals and/or deliver electrical stimulation in the brain. Currently, biodegradable and biocompatible materials have been actively investigated to create temporary electrodes that could degrade after serving their functions for neural recording and stimulation from days to months. The new class of biodegradable electrodes eliminate the necessity of secondary surgery for electrode removal. In this study, we created biodegradable, biocompatible, and implantable magnesium (Mg)-based microelectrodes for in vivo neural recording for the first time. Specifically, conductive poly-3,4-ethylenedioxythiophene (PEDOT) was first deposited onto Mg microwire substrates by electrochemical deposition, and a biodegradable insulating polymer was subsequently sprayed onto the surface of electrodes. The tip of electrodes was designed to be conductive for neural recording and stimulation, while the rest of electrodes was insulated with a polymer that is biocompatible with neural tissue. The charge storage capacity and impedance of Mg-based microelectrodes and their performance during neural recording in the auditory cortex of a mouse were studied. The results first demonstrated the capability of Mg-based microelectrodes for in vivo recording of multi-unit stimulus-evoked activity and spontaneous activity in the brain.

Small organic molecules exhibiting therapeutic properties are known as active pharmaceutical ingredients (API) and are of great interest in pharmaceutical material science. These API can exist in several crystalline forms as well as in different states of disorder of which the amorphous form is of great interest due to their greater free energy which results in higher bioavailability and exposure in-vivo. Detailed screening as well as subsequent characterization of the different forms with varying degrees of crystallinity is imperative to choosing a suitable lead candidate that will elicit the required therapeutic response in-vivo. Thus the material science aspect of drug development plays a vital role in the pharmaceutical industry since it is responsible for proper physical form selection and its maintenance in the formulation during its shelf life. Polymorphism, amorphization or changes to chemical composition are several ways in which the physical form of the API may be altered which may bring about a change in its pharmaceutical properties and ultimately affect the final drug product and its performance. This talk focuses on the material science part of pharmaceutical drug development and tackles the different challenges faced in characterizing drug substances in both the discovery and development stage as well as in the formulation. Judicious form screening as a salt or polymorph, form changes during processing and unit operations and vitrification of crystalline materials for improving bioavailability will be covered as well as analytical techniques used for such characterization. The focus is on several case studies involving both drug susbtance and drug product with emphasis on experimental (X-ray diffraction, spectroscopy, microscopy, thermal analysis) and computational characterization techniques to showcase advances in pharmaceutical material science encompassing both ordered and disordered materials.

Biosensors and materials for biomedical applications generally require precise chemical functionalization to bestow their surfaces with desired properties, such as specific molecular recognition and antifouling properties. Consequently, tailoring the chemistry at the biosensing interface has a crucial role in obtaining the best selectivity and sensitivity.¹ Especially for DNA biosensor, either biological or artificial probes, as well as antifouling moieties, need a defined type of chemistry to be anchored with respect to their chemical modification and the type of substrate, affecting the biodistribution.² In addition, control of the surface probe density is required for achieving high sensitivity.³ Traditional methods have the applicability limited to specific substrates and aim to control the density at the surface modification step, with the drawback of having to assess the hybridization efficiency each time.
Recently, polyelectrolytes were used in biosensing to tailor the probe type and distribution. Furthermore, the ensemble of substrates for biosensing purpose has been increased thanks to the multiple nature of polymers and their electrostatic interactions. The chemi/physisorption of modified polyelectrolytes provides advantages for the immobilization of biomolecules and for biosensing applications. At physiological pH, poly(L- lysine) (PLL) polymers readily and strongly adsorb onto a variety of metal oxide surfaces through multivalent electrostatic interactions between the positively charged lysine side-chains and a negatively charged surface.⁴ As a result, PLL polymers, which are easy to functionalize thanks to the amino groups in the side chain, allow the accommodation of the grafted functional moieties over the substrate, maintaining their adsorption properties.
Based on this approach, biorecognition surfaces were prepared by deposition of modified PLL polymers grafted with various fractions of oligo(ethylene glycol) (OEG, antifouling) and maleimide (Mal) moieties (PLL-OEG-Mal), so that both the type of functionalization and the control over the density is achieved at the same time, during the synthetic step, verified by ¹H-NMR. PLL-OEG-Mal polymers were self-assembled at the substrate and coupled to thiol-peptide nucleic acid (PNA) probes, forming a real-time DNA biosensor. A linear relationship between the probe density and the PLL grafting density was found monitoring the frequency shift of the hybridization step for the complementary DNA versus the density of Mal group coupled to the PLL backbone. In order to establish the absolute probe density values at the biosensor surfaces, cyclic voltammetry experiments using Methylene Blue-functionalized DNA were performed, providing a density of 1.24 × 10¹² probes per cm² per % of grafted Mal, thus confirming the validity of the density control in the synthetic PLL modification step without the need of further surface characterization.^5,6 Thanks to their advantages, modified PLL polymers can be used to tune the surface properties, accommodating several clickable side groups as linkers and antifouling moieties, and forming different architectures as layer-by-layer and µ-arrays, as well as their application in soft lithography and pillar structures.⁶

References:
1 D. Bizzotto, I. J. Burgess, T. Doneux, T. Sagara, H. Z. Yu, ACS Sensors 2018, 3, 5–12.
2 S. H. North, E. H. Lock, C. R. Taitt, S. G. Walton, Anal. Bioanal. Chem. 2010, 397, 925–933.
3 V. Biagiotti, A. Porchetta, S. Desiderati, K. W. Plaxco, G. Palleschi, F. Ricci, Anal. Bioanal. Chem. 2012, 402, 413–421.
4 S. Pasche, S. M. De Paul, J. Vörös, N. D. Spencer, M. Textor, Langmuir 2003, 19, 9216–9225.
5 J. Movilli, A. Rozzi, R. Ricciardi, R. Corradini, J. Huskens, Bioconjug. Chem. 2018, 29, 4110-4118.
6 J. Huskens, R. Ricciardi, J. Movilli, D. D. Iorio, A. Marti Morant, World Patent WO 2018/222034, 2018.

The oral cavity harbors a wide array of microbiota which if perturbed, would result in the loss of mutualistic/ symbiotic balance leading to diseases. The aggregates of these microorganisms can orderly lay in a protective sheath coined as extracellular polymeric substance (EPS), to form oral biofilm a.k.a. dental plaque. There is mounting evidence that the dental biofilm can be the culprit of several diseases such as dental caries which affects 2.4 billion people worldwide.
In the biofilm, the fermentation of the dietary carbohydrates causes organic acids production (pH~4.5) demineralizing the teeth enamel. The most common carious pathogen involved in the dental biofilm is the gram-positive bacteria, streptococcus mutans (S. mutans), which actively produces EPS and acids.
Unfortunately, the rigorous solution to the dental caries is compounded due to the multifactorial nature of the disease. Nanoparticles (NPs) can offer an unequivocal solution to the biofilm issue with the privilege of multifunctionality, on- demand controlled release of drugs, high loading efficiency, selectivity, and trackability. Importantly, the nanoparticles can be designed to be ‘self-contained’ and exert the therapeutic effects without the utilization of conventional antibiotics . Nevertheless, as a major group of nanobiotics, the experimentation with metallic nanoparticles has raised concerns regarding their accumulation and non-degradation.
Carbon dots (CDots) are the emergent class of carbon family which have attracted a host of research in recent years due to the ease of fabrication, tunable luminescent properties, and the abundance of functional groups. These properties combined with their degradability offer a unique platform for combating bacteria.
Herein, we present for the first time, a ‘particle in particle’ approach for targeting the EPS with its characteristic pH to release the load of inherently therapeutic CDots to kill notorious S. mutans. Specifically, phosphonium containing CDots have been wrapped in a layer of poly(styrene)-b-poly(n,n-dimethylaminoethyl methacrylate) (PS-b- PDMA) which shows pH responsiveness upon encountering acidic pH. Moreover, phosphonium ions have been utilized to confer antibacterial properties to the NPs as with more pronounced inhibitory effect compared to their ammonium counterparts.
The retained load of therapeutic CDots will get liberated only when in touch with the pH of EPS. This approach would not only lead to the biofilm dispersal and enhanced the bacterial susceptibility but would also exert the antibacterial properties in situ without any external drugs. Moreover, the absence of any external stimuli, for example, H2O2 and photostimulation, is a boon which reduces the unintentional toxicity.
We have demonstrated that these NPs could suppress the biofilm EPS and decrease the S. mutans viability in the in vitro human tooth model of the biofilm. The mechanism of action can be classified as ROS generation and fragmentation of genomic DNA on the subcellular level. We have shown, in vivo, in the rat model of dental biofilm that these nanoparticles could successfully decrease the cultivatable S mutans within 13 days using the plate count assay and S. mutans detection kit. It has been recognized through histopathological examination that these NPs did not interfere with the normal activities of the major organs while the dental caries scoring indicated a decrease in the caries development. The gene expression profiling of the oral microbiota from rats revealed that the change in the microbiome diversity was not statistically significant among groups. Therefore, our approach has great implication for the application of nanomaterials as degradable antibiofilm in the dental clinic.

Numerous bacterial stains have become resistant to conventional antibiotics in recent years. Fortunately, an increasing body of research indicates that through the addition of specific metabolites (like sugars), the antibacterial activity of certain drugs can be enhanced. A new type of self-assembled nano-peptide amphiphile (SANPA) was designed in this study to treat antibiotic resistant bacterial infections and to reduce the use of antibiotics. Here, SANPAs were self-assembled into nanorod structures with a nanoscale diameter at concentrations greater than the critical micelle concentration (CMC). Both Gram-positive and Gram-negative bacteria were treated with SANPAs with metabolites supplementation. After various amounts of time metabolites pre-incubation, SANPAs reduced bacteria growth relative to non-matebolite treatments at all concentrations. Cytotoxicity assays indicated that the presence of metabolites seemed to slightly ameliorate the cytotoxic effect of the treatment on model human fetal osteoblasts (or bone forming cells) and human dermal fibroblasts. In conclusion, we demonstrated here that SANPAs-like nanomaterials have a promising potential to treat antibiotic resistant bacteria especially when added to metabolites, potentially limiting their associated infections. By comparision of SANPAs treatments under different communication conditions, the results further provide resource for researcher to design novel agents and treatment methods against antibiotic resistant bacterial infection.

The study of the intracellular compartment requires devices that can not only monitor the bioelectric activity, but also control and observe the biochemical environment at the biomolecular level. Up to now the electrical activity of excitable cells, in particular that of a whole cells network has been studied with multi electrode array (MEA) devices. To overcome the intrinsic limitations of this method, such as low accuracy and low signal to noise ratio, 3D nanoelectrodes were developed on the planar surface in order to improve the interface with cells.¹ In fact, it has been amply demonstrated the strong coupling between these structures and the cellular membrane². A novel fabrication technique for producing 3D vertical nanostructures that can be tuned in material and shape has recently been shown by our group³. This technique enables the fabrication of 3D hollow nanoantennas with high control over geometry and layout by means of a focused ion beam (FIB) procedure. These plasmonic vertical structures have shown to be suitable for enhancing the electrical recording of electrogenic cells, for performing selective intracellular drug delivery^4,5To further increase the capabilities of MEAs, here we combine these 3D hollow nanoantennas on top of MEA electrodes fabricated on thin silicon nitride membranes in order to combine the improved electrical recording with a controlled delivery over the cells. The goal of in vitro devices is to replicate as closely as possible aspects of the true in vivo microenvironment, this platform could be applied to make in vitro assays more realistic and capable of a precise manipulation of the microenvironment to deliver soluble factors to cells. The nanoantenna’s hollow shape, in fact, allows to have a controlled delivery of molecules during cell signal recordings, providing the device with great versatility and allowing the exploitation of different approaches and techniques such as disease diagnosis, single cell analysis, drug screening, proteomics and other biological applications. The device configuration could provide flexibility in controlling the critical biochemical factors that influence cells behavior, allow for the partial differentiation of cells in a single system, provide for the establishment of biochemical gradients in two- or three-dimensional cultures, and at the same time allows for high quality intra and extracellular recordings.
1. Spira, M. E. & Hai,. Nat. Nanotechnol. 8, 83–94 (2013).
2. Dipalo, M. et al. Nano Lett. 9, 6100-6105 (2018).
3. Dipalo, M. et al. Nanoscale 7, 3703–11 (2015).
4. Dipalo, M. et al. Nano Lett. 17, 3932–3939 (2017).
5. Caprettini, V. et al. Sci. Rep. 7, 8524 (2017).

More than 2 million End Stage Renal Disease (ESRD) patients receive dialysis to sustain life, with this number likely to represent less than 10% of the actual need. In the United States alone, over 460,000 people are on dialysis. Conventional hemodialysis removes urea and other metabolic waste from the body by running ~120 L of dialysate over hollow fiber dialysis membranes each session, which is typically 3-4 hours and 3 times a week. The intermittent character of hemodialysis results in large fluctuations in blood metabolite concentrations. Observations show that long-term survival in dialysis patients treated by extended hemodialysis are improved compared to conventional hemodialysis. Thus a portable dialysis machine that is working continuously would bring in significant health, quality of life and economic benefits.

To enable portable kidney dialysis for ESRD, the regeneration of the dialysate in a closed loop system is a primary critical technical barrier. Currently the ~120 L (kg) of dialysate per session, far exceeds usable weights for a portable system. We have developed an efficient photooxidation system based on hydrothermally grown TiO₂ nanowires, UV LEDs, and catalytic gas diffusion barriers to decompose urea from the dialysate at rates sufficient to remove daily production of urea at 15 g/day.

A photoelectrical decomposition cell with SiO₂/FTO/TiO₂-nanowire anode, 10 mM urea/0.15 M NaCl electrolyte, and 4 mg/cm² Pt black loaded carbon paper cathode was characterized for urea decomposition efficiency. Under 4 mW/cm² illumination of the 365 nm LED with 40% quantum efficiency, the device yielded a photocurrent density of ~ 1 mA/cm² in a dialysate simulant of corresponding to 40% quantum efficiency in urea decomposition per incident photon. From performance parameters, a feasible portable device with ~0.23 m² active area and a current draw of 11 A is able to decompose a daily 15 g urea production sufficient to regenerate dialysate. Also high selectivity (80%) for urea decomposition over Cl₂ formation is shown. For comparison, prior reports in literature [J. Photochem. Photobiol. A-Chem, 2009, 205, 168] for urea fuel cells of agricultural waste, > 5000 A would be required. Improvements reported here are based on nanowire microstructure to efficiently separate electron holed pairs and efficiently adsorb incident UV irradiation.

Recent advances in molecular biology, single cell manipulation and data analysis techniques have allowed us to look into the molecular states of individual cells with unprecedented detail. These methods have made it possible to investigate how the genotype in combination with internal and external regulatory factors guide the development of complex and diverse phenotype. However, these methods rely on cell lysis and can provide only a snapshot of the cellular state. This makes it challenging to monitor gene expression changes over time. Inferences about dynamic cellular processes are obtained by constructing pseudo-time trajectories from the continuum of molecular states present in a cell population that can reflect the essence of a single cell trajectory. However, the dynamics of cell state may be non-hierarchical or stochastic which the computational algorithms cannot always capture. In order to acquire a holistic understanding of decision-making pathways involved in processes such as cell reprogramming, differentiation and maturation, tracking the same cell in time is necessary. Our long-term goal is to develop a single-cell microfluidics platform that can temporally examine cells by performing precise cellular manipulations (e.g. genetic editing using CRISPR/Cas9) and subsequent analysis of internal cellular biomarkers in a non-destructive manner. Such a system would allow us to monitor the biochemical changes in cells over time and correlate these to the external inputs and perturbations.
Localized electroporation has emerged as an effective technique for introducing the desired changes in cellular systems by delivering foreign molecules such as nucleic acids. Unlike bulk electroporation where the entire cell membrane is exposed to a strong and non-homogeneous electric field, localized electroporation utilized nanostructures (such as nanochannels) to confine the electric field to only a fraction of the plasma membrane. This controlled perturbation allows for efficient transport of molecules into the cells as well as helps maintain high cell viability. As such, localized electroporation is also being used to address the converse problem, i.e. to non-destructively sample the cytosolic contents of living cells.
Although several experimental reports demonstrate the phenomena of localized electroporation, a mechanistic understanding of the different parameters involved in the process is lacking. In this work, we present a multiphysics model that 1) estimates the transmembrane potential developed across the cell membrane in response to a localized electric field, 2) predicts the electro-pore distribution in response to the local transmembrane potential drop and 3) calculates the molecular transport into and out of the cell based on the predicted pore-sizes. Using the model, we identify that cell membrane tension plays a crucial role in enhancing both the amount and the uniformity of molecular transport, particularly for large proteins and plasmids. We also find that a critical voltage range is necessary for efficient electroporation. We qualitatively validate the model predictions by delivering large molecules (fluorescent-tagged bovine serum albumin and mCherry encoding plasmid) and by extracting an exogenous protein (tdTomato) in an engineered cell line on a localized electroporation platform. The findings presented here should inform the future design of microfluidic devices for localized electroporation based sampling and temporal, single cell analysis. Eventually, high throughput temporal analysis of single cells would help us gain insights into fundamental aspects of developmental biology, study the progression of diseases such as Alzheimer’s and Parkinson’s and evaluate the efficacy of drugs over time.

Endogenous electrical fields play a major role in various biological processes from embryonic development to cell division and migration (electrotaxis). The latter has been extensively researched due to its involvement in wound healing processes. A better understanding of electrotaxis induced by external electrical fields (direct current), could hence lead to a breakthrough in wound healing therapy. However, the underlying mechanisms within the cells during electrotaxis remain still unclear.
The main limitation for research, as well as the clinical application of direct current therapy for directed cell migration, lies within the utilized electrodes. Metal electrodes made of platinum or copper, as well as silver chloride, corrode and release toxic by-products under direct currents, which are harmful to cells and in addition, might influence their migratory function. Therefore, all electrotaxis experiments until now require agar salt bridges to connect the electrodes to the cell chamber, which lead to a bulky and inefficient experimental setup. A better solution for the stimulation material could improve cellular research and facilitate the transition towards clinical use.

As an alternative, we propose the use of polyimide-based iridium oxide electrodes coated with poly(3,4-ethylenedioxythiophene) polystyrene sulfonate (PEDOT/PSS). These electrodes are able to maintain stable ionic currents over long periods of time and can drive electromigration of cells without the release of any cytotoxic by-products.

Using a simple electrotactic chip we were able to show that PEDOT/PSS electrodes have sufficient ion filling capacity to drive electromigration of skin cells in an agar bridge free system. The chip was fabricated through soft lithography of polydimethylsiloxane (PDMS) which was chemically bonded to a glass coverslip through surface modification with oxygen plasma. The chamber consists of a channel with a seeding area for the cells and two reservoirs at both ends of the channel for medium and the insertion of the electrodes. The use of polyimide-based electrodes allows the reusability of these for several experiments, facilitates the assembly of the electrotactic chamber and greatly reduces the associated cost.
During the experiments, a constant voltage was applied to the electrodes and the current was continuously monitored. Time-lapse photography documented the directed migration and the cell motility was quantified utilizing circular statistics. It was found that pre-oxidation/reduction of the PEDOT/PSS electrodes increased the current injection and subsequently the possible charge delivery to the cells.
We conclude that PEDOT based electrodes can be efficiently used for direct current stimulation and have the power to influence the migration of cells relevant to the healing of human skin. The combination of polyimide-based reusable electrodes and disposable microfluidics make a cost-efficient analysis of electrotaxis possible at high throughput.

This project has received funding from the European Research Council (ERC) under the European Union’s Horizon 2020 research and innovation program (grant agreement No. 759655).

Dissolved oxygen [DO] is crucial to environment, industry, life technology, and human health, etc. Hypoxia[1] (≦0.5% O₂, usually at tumor environments) is relevant to cancer, stroke, arteriosclerosis, Parkinsonism, and Alzheimer's disease etc.. Therefore, in vivo dissolved oxygen detection is essential for disease prevention and treatment. For dissolved oxygen detection, phosphorescence based optical oxygen analysis showed superior performance among several ways with noninvasive sensing, no oxygen consumption, high sensitivity, fast responses and single cell scale sensing.
Herein, five kinds of multi-arm block copolymers were synthesized by atom transfer radical polymerization (ATRP) to load the excellent hydrophobic oxygen probe platinum (II)-5,10,15,20-tetrakis- (2,3,4,5,6-pentafluorophenyl)-porphyrin (PtTFPP) for optimizing PtTFPP's performance in hypoxia sensing and imaging. Among them, fluoropolymers were introduced with their excellent capacity to dissolve and carry oxygen for investigating carriers' structure-property relationship. Under nitrogen atmosphere, high quantum efficiency of PtTFPP in fluorine-containing micelles could reach to 22% without complicated modification of PtTFPP. Oxygen-nitrogen titration by a gas manipulator showed that fluorine-containing micelles have higher oxygen sensitivity (I₀/I) than fluorine-free ones, and micellar sensors are more sensitive under body temperature (37°C) than room temperature (25°C). Besides, the phosphorescence lifetime of fluorine-containing micelles achieved 76μs under nitrogen, which is among the longest ones in the known literature. This result showed great potential for lifetime sensing and imaging because longer phosphorescence lifetime determines accurate resolution and legible hypoxia imaging.
For biological application, one fluorine-containing PtTFPP micellar sensor was used to detect E.coli JM109's respiration under oil seal by a high throughput plate reader. Based on the Stern-Volmer equation and the dynamic phosphorescence intensity curve, we converted the phosphorescence intensity at each time point to the real-time DO concentration. We also extended the sensor's respiration application to mammalian cells by using J774A.1, MTT assay and confocal imaging were carried out for demonstrating the micellar sensor is nontoxic and extracellular. Similar to the phenomenon of E.coli, higher cell numbers induced faster oxygen consumption. In vivo hypoxia imaging of tumor-bearing mice was also achieved through an In Vivo Imaging System. The phosphorescence imaging was obtained after anesthesia a sensor injected mouse. It can be seen that the phosphorescence image at tumor region is much brighter, and the phosphorescence intensity at tumor region is 1.6 times higher than that of normal region. Moreover, the phosphorescence imaging lasted for at least 10 minutes without obvious decay, indicating that the PtTFPP possessed excellent in vivo stability and strong phosphorescence signal after being encapsulated by our materials. Thus, the sensors reported here might be capable for in vivo tumor imaging. It is expected that the further use of the sensors can be extended to measure the intravascular oxygen contents, which will be more instructive to cancer prevention and cancer clinical analysis.
Reference:
1. Vaupel, P.; Hoeckel, M.; Mayer, A. Detection and characterization of tumor hypoxia using pO2 histography. Antioxid. Redox Signal. 2007, 9, 1221-1235.

Human skin exhibits high stiffness of up to 100 MPa and high toughness of up to 3,600 J m^-2 despite its high water content of 40~70 wt%. Engineering hydrogels have rarely possessed both high stiffness and toughness, because compliant hydrogels usually become brittle when excess crosslinker is added to make the gel stiff. Furthermore, conventional hydrogels usually swell under physiological conditions, weakening their mechanical properties. Here, we designed a non-swellable hydrogel with high stiffness and toughness by interpenetrating covalently and ionically crosslinked networks. The stiffness is enhanced by utilizing ionic crosslinking sites fully, and the toughness is enhanced by adopting synergistic effects between energy-dissipation by ionic networks and crack-bridging by covalent networks. Non-swelling behaviors of the gel are achieved by densifying covalent and ionic crosslinks. The hybrid gel shows high elastic moduli (up to 108 MPa) and high fracture energies (up to 8,850 J m^-2). In vitro and in vivo swelling tests prove non-swelling behaviors of the gel. Live/dead assays show 99% cell viability over a period of 60 days.

Obesity is a global health issue that is suffered by over 700 million people worldwide.[1] Common approaches for treating obesity include non-surgical (excise) and surgical (invasive) treatments, which normally have a high potential of weight rebound or can introduce serious complications.[2] Recent studies demonstrated that vagus nerve blocking and stimulation have multiple physiologic functions related to food intake, energy metabolism, and glycemic control, which can result in a meaningful weight loss. Nevertheless, like many other electrical nerve
stimulation therapies, the electrical signals and corresponding biological activities are very hard to be correlated, particularly for such irregular food taking and stomach peristalsis activities. Therefore, the long-term efficacy may be jeopardized and the constant electrical stimulation may trigger compensation mechanisms.[3] Power supply is also an issue for this type of implanted electronic devices.
Here we present an implanted vagus nerve stimulation (VNS) system that is battery-free and spontaneously responsive to stomach movement. The VNS system comprises a flexible and biocompatible nanogenerator that is attached on the surface of stomach. It generates biphasic electric pulses in response to the peristalsis of stomach. The simulated satiety signals deceive the brain via vagal afferent fibres,[4] and reduces food intake when the stomach is in motion. This strategy was successfully demonstrated on rat models. Within 100 days, the average body weight was controlled at 350 g, 38% less than the control groups. This new VNS system correlated nerve stimulation with targeted organ functionality through a smart, self-responsive device, and demonstrated an outstanding weight control capability better than other electric stimulation strategies. This work also provides a new concept in therapeutic technology using artificial nerve signal generated from coordinated body activities.
References:
[1] Collaborators, G. O. et al. Health Effects of Overweight and Obesity in 195 Countries over 25 Years. N. ENGL. J. MED. 377, 13 (2017).
[2] Bray, G. A. & Tartaglia, L. A. Medicinal strategies in the treatment of obesity. Nature 404, 672-677 (2000).
[3] Schachter, S. C. & Saper, C. B. Progress in Epilepsy Research : Vagus Nerve Stimulation. Epilepsia 39 (1998).
[4] Schwartz, M. W., Woods, S. C., Porte, D., Jr, Seeley, R. J. & Baskin, D. G. Central nervous system control of food intake. Nature 404, 661 (2000

This talk summarizes concepts in materials science that serve as the basis for transient electronic implants, as a new paradigm for treating injuries and tracking recovery. Here, engineered device platforms provide monitoring and/or therapeutic functionality during the time course of a natural biological process such as healing, and then disappear into the body, without a trace, through mechanisms of bioresorption. This mode of operation eliminates unnecessary device load on the body, and associated risk to the patient, in a way that bypasses the need for secondary surgical extraction. Examples of bioresorbable materials and device designs will be presented in the context of (1) electrical stimulators for accelerated neuroregeneration in damaged peripheral nerves, (2) cardiac pacemakers for recovery from heart surgery and (3) programmable drug release vehicles for treatment of disease.

TiO₂ nanotube arrays constitute highly versatile scaffolds that are suited for long-term organotypic culture of neuronal tissue, including retina explants and beyond. While tube diameter and surface roughness [1] are central parameters for successful culture and have to be optimized for every specific tissue type, super hydrophilicity ensures nutrition supply by a wetting layer of culture medium even in absence of complicated perfusion systems [2]. Clearly adhesion between tissue explants and nanotube scaffolds plays a key role for maintaining tissue integrity. Lift-off approaches are, on the other hand, desirable for tissue transfer and nanotube regeneration, respectively. Employing extensive environmental scanning electron microscopy (ESEM) as well as laser scanning microscopy (LSM) studies we address both aspects and demonstrate that UV-light exposure and enzymatic treatment are ideally suitable for nanotube regeneration. Based on these findings, we present an assay for concurrent biomechanical straining and in situ imaging employing ESEM and LSM. Thus, we will be able to find correlations between structural components of retina and their variation in tissue mechanics. This will pave the way for in vitro studies on tissue regeneration and surgery techniques in combination with drug testing.
[1] K. Fischer, S. G. Mayr. Adv. Mater. 23, 3838–3841 (2011)
[2] V. Dallacasagrande, M. Zink, S. Huth, A. Jakob, M. Müller, A. Reichenbach, J.A. Käs, S.G. Mayr, Adv. Mater. 24, 2399–2403 (2012)

The study of the restoration of vision is one of the most popular research topics in neuroscience and biomedical sciences. A functional/adaptive retinal prosthesis is highly sought after to combat degenerative eye diseases and build foundational devices for future engineering models. A sub-retinal device requires a specific structure compared to other prosthesis. However, the specifics of this work also require high transparency as a performance metric. Low impedance and high Charge Injection Capacity (CIC) are typical metrics for stimulus electrodes but our chip architecture will place electrode materials anterior to photodiodes, thus a material with high transparency and biocompatibility must be determined.
These electrode characteristics have already been studied extensively in photovoltaics. Therefore in order to provide the best performance for our proposed device it is necessary to extensively test a selection of materials from previous literature. Each of the three categories: Trans-conductive Oxides(TCO), Metal Nanowires(NW), & 2-D materials can yield suitable candidates, such as: Aluminum doped Zinc Oxide (AZO), Boron doped Zinc Oxide (BZO), Indium Tin Oxide (ITO), AgNW(Silver Nanowires) + PEDOT:PSS, Carbon Nanotubes(CNT) + PEDOT:PSS, & Graphene with PEDOT:PSS. The primary materials examined in this study are Aluminum doped Zinc Oxide (AZO), Boron doped Zinc Oxide (BZO), PEDOT:PSS, and AgNW.
The fabrication of test electrodes has two separate wafer designs. The testing substrate used for electro-characteristics has a Ti/Au/Ti sandwich structure on Si wafers; on a SiO2 layer 30nm Ti was sputtered, followed by 500nm Au, and then 30nm or Ti. A photoresist pattern was used with wet etching to create a desired wiring pattern. Next, a 1um SiO2 passivation layer was applied and reserved contact/electrode locations were etched with RIE. Last, an experimental electrode material was sputtered onto the reserved electrode location. For the biocompatibility test and the transparency test, electrode material was sputtered onto 1mm thick quartz glass substrates. The resulting substrate was cut to specifications via optimized saw dicing. Optical testing utilizes the same substrate without dicing measures. Since our device will be mounted in ocular tissue for the long-term biocompatibility concerns become a central issue. An MTS biocompatibility assay test, using Hippocampal neurocytes, was performed on 10mm x 10mm square test substrates on quartz glass.
AZO, BZO, & PEDOT:PSS electrodes were successfully formed and Pt and Ti electrode wafers were fabricated for baseline. In the biocompatibility test. Each conductive material is deposited on quartz pillars. The pillars have dimensions of 50µm x 50µm x 200µm. Each pillar is spaced 200µm X & Y from other pillars in a grid like fashion. This research currently is verifying the bio-compatibility of flagged materials and we will continue to report on electrode performance compared to referenced research. AZO was a promising material but with initial testing was shown to be incompatible. Statistically speaking AZO is less biocompatible than control condition so is incompatible with our application at this point in time.
BZO and AgNW are also being targeted for Biocompatibility testing due to the potential of harmful nanoparticles from the electrode material. PEDOT:PSS is commended for its biocompatibility, electrical, & optical characteristics but process fabrications methods used can lead to undesirable material conditions like optical hazing in the polymer.
The Hippocampal neurocytes in direct contact with our electrode material showed biocompatibility of AZO to be lower than Pt electrodes. Further testing with BZO and AgNW are required because current literature suggests that each of the metallic materials has the potential to release hazardous nanoparticles to the tissue. Optical and electric characteristics have thus far been within expected performance ranges for our desired device.

Hyperthermia (also known as thermal therapy or thermal ablation) has been adopted as one of the cancer treatment approaches in medical surgery. By applying high temperature on the cells, the proteins structure would be damaged and the cell would be destroyed. In general, Radio Frequency (RF) probes are applied onto the target tissues for the ablation process. However, one of the major drawback is difficult to apply when the cancer tissues are on the surface of the organs. Here we propose a novel approach to develop surface hyperthermia technique by using oscillating magnetic field induction on metal thin film. In our device, the heat is transferred from the organ surface to the cancer cells through bioheat transfer process. Heat generation process will be modulated by varying the conductance of the metal thin film, the strength and the frequency of the oscillating magnetic field. Optimal parameters such as film geometry, film thickness, magnetic induction distance and ablation time are identified through the combination of ex vivo experiment and computational modelling. Near-field communication (NFC) temperature sensor is also integrated in the system enabling a wireless interfaces for smartphone to monitor and analyze the treatment performance. Based on the steady state heat transfer model and comparing the measured temperatures by the NFC temperature sensor, we can calculate the temperature profile inside the pig liver. By using a input power of 2000W/m², the highest heater temperature is 443 K in our device which is sufficient for the ablation process. This wireless heater is believed to have high potentials in various kind of in-vivo testing or non-invasive thermal treatments.

Tight interaction between nanophysics, materials science and biotechnology led to the emergence of a new class of bioinspired systems that enables to bring the area of biosensorics e.g. for cell or molecular diagnostics and analytics to the new level. Very promising candidates for the future diagnostics are the electronic nanobiosensors that have attracted great attention in the last years since they provide rich quantitative information for medical and biological assays without pre-treatment and specific optical labelling of the detected analyte. One dimensional nanostructures, e.g. semiconductor and metallic nanowires serving as backbone of the sensor device, have attracted attention as highly efficient elements due to their high surface-to-volume ratio, which simplifies the detection of biochemical species. Use of nanowires enable to ultimately decrease the dimensions of the sensing area of the device and thus increase the resulting sensitivity of the assay. Finally, nanoscale sensors integrated into lab-on-a-chip system offer attractive opportunity of the multifunctional and multiplexed bio- and chemical analysis that can be performed in real time, directly in-flow.
Here we focus on two subsystems for the in-flow analysis at the micro- and nanoscale, represented by (a) silicon nanowires based field effect transistor and (b) metal nanowires assembled as nanocapacitor. We demonstrate the applicability of the systems for the detection single molecules, e.g. influenza or Ebola viruses [1-3], biochemical reactions [4], as well as classify the blood cells in a cytometry format.

[1] D. Karnaushenko et al. Adv. Health. Mater. 4 (10), 1517-1525 (2015).
[2] M. Medina-Sanchez et al. Nano Letters 16 (7), 4288-4296 (2016).
[3] B. Ibarlucea et al. Nano Research 11 (2), 1057-1068 (2018).
[4] J. Schütt et al. Nano Letters 16 (8), 4991-5000 (2016).

Highly sensitive wearable sensors that can be conformably attached to human skin or integrated with textiles to monitor the physiological parameters of human body or the surrounding environment have garnered tremendous interest. Owing to the large surface area and outstanding material properties, nanomaterials are promising building blocks for wearable sensors. In this talk I will start with a brief overview of the recent advances in the nanomaterial-enabled wearable sensors including temperature, electrophysiological, strain, tactile, electrochemical, and environmental sensors. Integration of multiple sensors for multimodal sensing and integration with other components into wearable systems are summarized. Then I will present the array of wearable sensors developed in my group using silver nanowire (AgNW) composites. AgNW composites offer outstanding combination of high conductivity and high stretchability. Finally I will discuss a major challenge in the field of nano-enabled wearable sensors – scalable nanomanufacturing – and some recent work addressing the challenge.

In 2018, an estimated 266,120 new cases of breast cancer (BC) will be diagnosed in the United States, and 40,920 cases will die from the disease mainly due to metastasis. For example, according to the American Cancer Society, while the survival rates of BC stages 0-1 are approximately 100% and 93% respectively, metastasized BC has only a 22% survival rate. Understanding circulating tumor cells (CTCs) present a valuable opportunity for possible strategies to reduce dissemination. Currently, the only FDA-approved system (CELLSEARCH^®) enumerates merely a total number of chemically-fixed CTCs and cannot capture CTC clusters.
Microfluidics is an active field of research for isolation of CTCs. Microfluidic technologies such as CTC-Chip, Herringbone chip, CTC-iChip, Vortex, Rarecyte, Fluxion, NanoVelcro, DEP-Array, and JETTA are leading fluidic devices. There are about 400 different microfluidic techniques reported in the literature for CTC capture since 2007. Although promising, there are inherent constraints in large scale production of complex microfluidic based devices and surface functionalization for targeted capture. Further, the rare CTC clusters could potentially be damaged in fluidic devices due to turbulence, vortices and shear forces. Very few devices have been able to successfully capture CTC clusters with 30-40% sensitivity in metastatic patients. Further, some of the leading microfluidic technology has only 5-44% purity which makes genomic sequencing difficult.
By engineering nanomaterials namely carbon nanotubes, and microarray manufacturing techniques, we have been able to successfully develop the world’s first microarray for CTC capture. Advantages of our microarrays include: 1) microarray format enabling large volume of blood to be fractionated into smaller portion that enables better capture sensitivity; 2) variety of antibodies functionalized in the same array, resulting in CTC capture based on multiple markers from a single blood sample; 3) no transfer of cells is necessary to do microscopy, and one can do confocal/optical microscopy on chip; 4) surface architecture lends itself to variety of surface functionalization and instant CTC isolation for further analysis unlike microfluidics where CTCs have to be recovered from sealed chambers and could be lost; and 5) mature manufacturing process resulting in a 99% yield.
Using the mentioned advantages of microarrays, we have demonstrated the capture of spiked breast cancer cells in blood using both antigen-dependent and independent capture with >90% capture rate and elimination of 99% contaminating leukocytes. The microarray devices are electrically conductive, that enables electrical signals and mathematical classification techniques to capture breast cancer SKBR3 cells in blood with 90% sensitivity and 90% specificity. Further, using triple negative breast cancer patient derived xenografts (PDX), we have demonstrated the capture of highly invasive CTCs, CTCs with microtentacles, and the very rare CTC clusters which are metastasis initiating cells. In volumes up to 0.8 ml blood from PDX-mice, we have shown the capture of up to 26 CTC clusters and CTCs of different morphologies. The combination of materials engineering, microarray manufacturing, biochemical surface functionalization and cancer biology has led to this new microarray technology that will help in fight against cancer, the results of which will be presented at the spring 2018 MRS conference.

Autoimmune disorder is a kind of common chronic disease, which is difficult to cure throughout life. Small chemical drugs such as glucocorticoids, immunosuppressors and non-steroidal anti-inflammatory drugs have been widely used for the treatment of autoimmune disorders. However, these small chemical drugs suffer from poor solubility, short circulating half-life and adverse side effects, which lead to poor compliance to patients and limit the widely clinical use. One of the most effective strategies to extend the circulating time is loading drugs into nanocarriers to form nanomedicines, which is of particular interest for cancer and viral diseases therapy but seldom applied in autoimmune disorder treatment. Furthermore, current carriers have many drawbacks such as poor biocompatibility, low stability and over-complicated design. In this study, we developed an easy but general drug delivery platform based on the new polyhydroxyalkanoate terpolymer-poly(3-hydroxybutyrate-co-3-hydroxyvalerate-co-3-hydroxyhexanoate) (PHBVHHx). We reported the first example of PHBVHHx nanoparticle loaded non-steroidal anti-inflammatory drug, ultimately being applied in systemic lupus erythematosus therapy. These nanoparticle are biodegradable, stable, and show improved pharmacokinetics, optimized biodistribution, low systemic toxicity and excellent in vivo therapeutic efficacy in a MLR/lpr murine model of systemic lupus erythematosus. This delivery system may provide a new and general platform for the development of nanomedicines with enhanced therapeutic efficacy and reduced side effects.

Antimicrobial resistance is a global health crisis that complicates the prevention and treatment of diseases caused by pathogenic microbes. In particular, the formation of bacterial biofilms on medical implants protects the pathogen from the host’s immune response and antibiotic treatment, causing chronic infections that form the basis of persistent microbial infections in the human body. To date, the treatment of medical device related infections due to biofilms remains a huge challenge in the clinical practice setting, mainly due to the extreme difficulty in the disruption and eradication of bacterial biofilm growth, which is resistant to antibiotics as well as mechanical treatment approaches. Recent advancements on the development of smart hydrogel nanocomposite systems with tailored properties provide promising routes to develop novel treatments that can be used to disrupt and completely eradicate pre-formed biofilms. In this talk, we will present our work on the development of an externally actuated thermoresponsive hydrogel nanocomposite system, containing D-amino acids (D-AAs) and engineered nanoparticles, which can be delivered and transforms from a solution to a gel state at physiological temperature for sustained release of D-AAs and light or magnetic field-actuated thermal treatment of targeted infection sites. The D-AAs in the nanocomposite are known to inhibit biofilm formation and also disrupt existing biofilms. In addition, loading the hydrogel nanocomposite with magnetic or gold nanoparticles allows for combination thermal treatment following magnetic field or light stimulation, respectively. Using this novel two-step approach to utilize an externally actuated hydrogel nanocomposite system for thermal treatment, following initial disruption with D-AAs, we were able to successfully demonstrate the effective disruption and total eradication of Staphylococcus aureus biofilms, which were resistant to conventional antibiotics used in the clinic and were not completely cleared using individual D-AA or thermal treatments.

Rapid progress in nanotechnology allows us to develop a large number of innovative wearables such as activity trackers, advanced textiles, and healthcare devices. However, manufacturing processes for desirable nanostructure are usually complex and expensive. Moreover, materials used for these devices are mainly derived from nonrenewable resources. Therefore, it poses growing problems for the living environment, and causes incompatible discomfort for human beings with long–time wearing. Here, a self–powered cellulose fiber-based triboelectric nanogenerator (cf–TENG) system is presented through developing 1D eco–friendly cellulose microfibers/nanofibers (CMFs/CNFs) into 2D CMFs/CNFs/Ag hierarchical nanostructure. Silver nanofibers membrane is successfully introduced into the cf–TENG system by using CMFs/CNFs as the template, which shows excellent antibacterial activity. Enabled by its desirable porous nanostructure and unique electricity generation feature, the cf–TENG system is capable of removing PM_2.5 with high efficiency of 98.83% and monitoring breathing status without using an external power supply. This work provides a novel and sustainable strategy for self–powered wearable electronics in healthcare applications, and furthermore paves a way for next–generation flexible, biocompatible electronics.

Carbon dots, a new class of zero-dimensional carbon-based nanomaterials, have attracted great interest recently in the nanomedicine because of their versatile merits such as biocompatibility, facile functionalization, and tunable photoluminescence properties. In particular, carbon dots have allowed for their biomedical applications to disease diagnostics and therapeutics such as bioimaging and photodynamic therapy. Here, we present the new design of carbon dots with multiple functions that highly inhibit the aggregation of Alzheimer’s amyloid-β (Aβ) peptides.
The abnormal assembly and aggregation of Aβ peptides in the brain tissue is a major hallmark of Alzheimer’s diseases (AD) that currently affects one in nine people aged over 65 worldwide. Despite numerous efforts made over the past decades, effective suppression of Aβ aggregation is still challenging due to multifactorial pathogenesis involving high levels of metal ions in the brain. Note that copper ions (Cu(II)) have drawn significant attention in AD pathogenesis due to their high activities in binding with Aβ peptides and forming neurotoxic Aβ-Cu(II) complexes.
To provide a new anti-amyloidogenic strategy in the prevention of Cu(II)-associated Aβ aggregation, we have synthesized the multifunctional carbon dots and demonstrated their capabilities on (i) effective Cu(II) coordination, (ii) interruption of Aβ self-assembly, and (iii) photodynamic oxygenation of Aβ residues. The nitrogen-containing aromatic moieties on as-prepared carbon dot’s surface do a key role in interact with Cu(II) and Aβ’s aggregation sites by electrostatic and hydrogen/hydrophobic interactions. Interestingly, the Cu(II) coordination of carbon dots promotes the carbon dots’ absorption and fluorescence enhancement, thus further leads the photoinduced generation of reactive oxygen species (ROS) under light irradiation; the resulting ROS from carbon dots then photodynamically oxygenated Aβ residues (e.g., histidine and methionine) to lose the affinity towards Cu(II) as well as an aggregative property. These carbon dots’ simultaneous and multiple inhibitory effects significantly mitigated the neuronal cytotoxicity of Cu(II)-bound Aβ species, showing 96% of cell survival rate.
In summary, we will introduce the synthesis of multifunctional carbon dots and the various photochemical analyses to validate the suppressing capabilities of carbon dots against Aβ aggregation. Our investigations highlight the distinctive surface and luminescence properties of carbon dots as promising biocompatible nanoagents and their therapeutic application in preventing Aβ-induced Alzheimer’s diseases.

The Nobel Prize in Chemistry 1987 was awarded to Cram, Lehn and Pedersen for their development and use of molecules with structure-specific interactions of high selectivity. Twenty years later in 2016, Nobel Prize in Chemistry recognized supramolecular chemistry and molecular recognition again by awarding Sauvage, Stoddart and Feringa for the design and synthesis of molecular machines. This talk takes cyclodextrin and cucurbituril as examples to develop cancer nanotheranostics that make use of the dynamic and responsive nature of non-covalent interactions. By the formation of host–guest complexes with macrocyclic hosts, the anticancer drugs can be easily formulated to prepare nanomedicines showing satisfactory anti-tumor efficacy and reduced normal organ toxicity.

Collagen (Col) is a well-studied biomaterial, particularly as wound dressing material due to its superior biological and chemical properties [1]. However, the application of collagen in wound healing remains as a critical issue due to poor adhesion, proliferation and migration of cells. In addition, majority of the collagen-based biomaterials are primarily fabricated from mammalian sources, such as bovine and porcine [2]. There have been many considerations in terms of religious issues. Moreover, the extraction process from mammalian animals is complex, laborious, time consuming and expensive due to the stiff and fibrous nature of bovine and porcine tissue [3]. Hence, alternative sources of collagen are highly desirable, particularly for wounds treatment.
In this study, collagen derived from the skin of the American Bullfrog (i.e. Rana catesbeiana) is explored as nature-derived biomaterials for wound healing applications. We report the successful implementation of an innovative method that can significantly shorten the collagen extraction process by ~50% compared to traditional acid solubilisation method with relatively high extraction yield (~35%). Differential scanning calorimetry (DSC) analysis suggest that the collagen has excellent thermal stability (~41°C), suggesting that it is compatible for skin-contacting as well as for in vivo implantation applications. In vitro studies showed that the bioactivity of the bullfrog skin-derived collagen was well-preserved by supporting the attachment and proliferation of human keratinocytes (i.e. HaCaT), compared to commercially-available bovine collagen. Interestingly, our findings demonstrated that the extracted bullfrog skin-derived collagen can significantly enhanced the wound closure rate of HaCaT in an in vitro scratch wound assay as compared to bovine collagen. At the mechanistic level, we showed that this phenomenon may be attributed to the ultra-fine nano-fibers of the frog-skin derived collagen, which could augment the process of re-epithelization by modulating the adhesiveness and chemotaxis of the keratinocytes [4]. Overall, bullfrog skin could be a valuable waste-to-resource material for the production of non-mammalian collagen with high yield percentage and great wound healing capacity.
References
[1] Chattopadhyay, S. and Raines, R.T., 2014. Review collagen-based biomaterials for wound healing. Biopolymers, 101(8), pp.821-833.
[2] Gould, L.J., 2016. Topical collagen-based biomaterials for chronic wounds: rationale and clinical application. Advances in wound care, 5(1), pp.19-31.
[3] Gorlov, I.F., Titov, E.I., Semenov, G.V., Slozhenkina, M.I., Sokolov, A.Y., Omarov, R.S., Goncharov, A.I., Zlobina, E.Y., Litvinova, E.V. and Karpenko, E.V., 2018. Collagen from porcine skin: a method of extraction and structural properties. International Journal of Food Properties, 21(1), pp.1031-1042.

The complexity and heterogeneity of cancer and subsequently developed multidrug resistance (MDR) seriously undermines the therapeutic potential of chemotherapy. Herein, we developed a biodegradable redox-heat assisted cancer killing (RHACK) nanoparticle for enhanced treatment of cancer in vitro and in vivo. Unlike conventional anticancer strategies, our hybrid nanoparticle based drug delivery system (DDS) not only target-specifically delivers anti-cancer reagents, but also targeting heterogeneous MDR pathways to achieve sensitization of cancer cells. More specifically, our RHACK nanoparticle-based drug delivery system (DDS) reduces intracellular glutathione (GSH) level, induces ROS generation and simultaneous provide photothermal (PTT), thereby synergistically sensitizing breast cancer towards chemothreapy. Remarkably, our RHACK nanoparticle is constructed from an iRGD conjugated hollow iron oxide carbon core and a manganese dioxide shell (Fe₃O₄-C@MnO₂-iRGD NPs) and shows unique benefits for synergistically overcoming MDR: i) a strong NIR absorption for effective laser ablation of tumor and sensitize the tumor to further chemotherapy; ii) a tunable glutathione-reactive MnO₂ shell; iii) a redox-based endogenous biodegradability; iv) a hollow structure for efficient anti-cancer drug loading; v) Fe³⁺ as degradation product for induction of intracellular Fenton reaction and more toxic ROS species (hydroxyl radical); vi) Mn²⁺ as the second degradation product for T1 MRI imaging; and vii) capabilities of breast tumor targeting and penetration. Based on this unique platform, we successfully demonstrate a robust co-sensitization of a malignant drug-resistant breast cancer towards a clinical applied anti-cancer drug in vitro and in vivo by combining PTT, glutathione reduction and Fenton reaction on a single platform. A detailed mechanistic study on the molecular biology pathways further reveals the synergy originate from the regulation over several key genes and proteins including HSF-1and MDR-1/P-gp, TP53, BCL-2, BAX, Caspase 3/9. Given the heterogenous tumor microenvironments and the highly complicated drug-resistance pathways in breast cancer, our multifunctional hybrid nanoparticle-based DDS could provide a promising therapeutic alternative for more efficient treatment of breast cancer in clinical applications.

Materials engineering is typically divided into “bottom-up” and “top-down” methodologies. While, top-down engineering has been common practice for centuries, these approaches are rarely used with respect to biological systems. We have applied a top-down approach for generating a mineral gradient scaffold in trabecular bone. Mineral gradients are found in a variety of biological systems that link soft tissue and bone. Some of these systems are primarily mechanical, such as the attachments of ligaments and tendons with bone or the interface between cartilage and bone. Other soft tissue-to-bone interfaces are necessary for development, such as the growth plate or the interface present during endochondral ossification. These interfaces are further relevant to cancer metastasis, as tumors have been found to localize to the growth plate. These systems are complex, requiring a method for producing the mineral gradient present at the interface. With these goals in mind, we fabricated a mineral gradient scaffold through the spatially controlled removal of mineral from bone.
To fabricate this scaffold, cylindrical trabecular bone biopsies were extracted from neonatal bovine femurs. These biopsies were decellularized, followed by partial submersion in a demineralizing solution. Scaffolds were removed from the demineralizing solution at time points up to 6 hours, finding that 4.5 hours represents the median point in the mineral removal process. The main factor driving the amount of mineral removed from the biopsy was observed to be the porosity in the trabecular bone. This observation was confirmed by plotting porosity vs. time vs. the fractional demineralized content and fitting a surface to these data. Scaffolds were segmented into quarters, and X-ray diffraction was performed on these segments, confirming the presence of a mineral gradient within the scaffold.
To examine the cellular compatibility of the resulting mineral gradient, mesenchymal stem cells were seeded onto these scaffolds. The seeded mineral gradient scaffolds were cultured in osteogenic media, and a live/dead stain was performed. The cells attached to the exterior of the trabecular structure with a viability of ~91%. The scaffolds exhibited high viability and with no differences in viability were observed in the mineralized or demineralized portions of the scaffolds.
Immunohistology was performed to examine the effects of mineral content on cellular behavior. Seeded scaffolds were cultured, fixed, and sectioned for staining. Stains were chosen to determine if stem cells were showing any chondrogenic or osteogenic behavior. Staining profiles revealed differences in cellular behavior between the mineralized and demineralized ends as well as on the exterior and interior of the scaffold. Type I collagen staining was observed throughout the scaffold. Type II collagen staining was consistent on the interior of the scaffold but appeared higher in the exterior regions of the mineralized end of the scaffold versus the demineralized end. Alkaline phosphatase staining was consistent on the exterior of the scaffold but appeared higher on the mineralized interior vs. the demineralized interior. These findings reveal that the presence of a mineral gradient drives stem cells to produce proteins associated with chondrogenic and osteogenic cellular behavior.
Overall, we fabricated mineral gradient scaffolds using a top-down engineering approach. These scaffolds, generated using a novel approach to biomaterials synthesis, drive cellular behavior and will be useful in a variety of fields, including orthopedics, development, and cancer research.

Amyotrophic lateral sclerosis (ALS) is the most common adult-onset motor neuron disease, characterized by a rapid loss of upper and lower motor-neurons resulting in patient death from respiratory failure within 3-5 years of initial symptoms onset. Although at least 30 genes of major effect have been reported, the pathobiology of ALS is not well understood. There is a critical need to develop a system which can accurately diagnose this rapidly deteriorating disease. Herein, we report on graphene’s phonon vibration-energies as a sensitive measure of the composite dipole moment of the components of the interfaced cerebrospinal fluid (CSF) to specifically identify patients with ALS disease. The second-order overtone of in-plane phonon vibration energy (2D) of graphene shifts by 3.2±0.5 cm^-1 for all ALS patients studied in this work. Further, the amount of n-doping induced shift in phonon energy of graphene, interfaced with CSF, is specific to the investigated neurodegenerative disease (ALS, Multiple Sclerosis and Motor Neuron Disease). By removing a severe roadblock in disease detection, this technology can be applied to study diagnostic biomarkers for researchers developing therapeutics and clinicians initiating treatments for neurodegenerative diseases.

Cells are continuously performing chemical computations based on a range of inputs from their immediate environment. Two fundamental phenotypes that result from these computations are cell adhesion and migration, which play central roles in development, wound healing and diseases such as cancer. The degree to which cells adhere or migrate is dependent upon a combination of chemical and physical properties of the substrate. However, the complex coupling amongst these physical and chemical cues has made it challenging experimentally to deconvolve their individual contributions. Furthermore, cells dynamically alter their environment by the secretion of proteins. To address these complexities, we have employed a combination of electron beam and optical lithographic techniques to fabricate substrates which incorporate independently tunable topographical and chemical signaling cues, as well as nanosensors for detecting cell secretions. The approach works by patterning two materials on a single chip, one for topography, and the other for chemical functionalization. The first material, quartz, is light microscopy compatible and can be dry etched for 3D topography formation. The second material, gold, is patterned independently of the quartz etching process into nanostructures for use in either interfacing with the cell membrane or detecting protein secretions. I will present results on a range of cell types cultured on multiplexed substrates that integrate nanosensors for quantifying single cell secretions, topographical patterns for directing cell migration and biofunctionalized nanostructures for targeted adhesion studies. Cells at the boundary of two patterns experience a surface-induced competition that can be tracked in real time as they decide to adhere to one surface or migrate to the other. These chips enable quantitative insight into the biology of how local environments influence cellular decisions as well as potential guidance in the design of next generation wound healing materials and implants.

The dynamic response of cells when subjected to mechanical impact has become increasingly relevant for accurate assessment of potential blunt injuries and understanding injury mechanisms. When exposed to a blast, ballistic, or impact, a biological system, e.g., human brain or skin, is rapidly accelerated, which results in an acceleration-induced pressure gradient. In order to study cell behavior under these threats, we have developed a new experimental method that applies a well-controlled mechanical impact to live cells cultured in a custom-built in vitro setup compatible with live cell microscopy. A drop tower system is utilized to apply the mechanical impact while concurrently visualizing each impact using high speed cameras. The impact amplitude is controlled by changing the vertical height of a movable mass, i.e., drop height, which is then accelerated toward the setup once released. Our experiments show that the maximum pressure in the in vitro setup is very sensitive to the height of cell culture media. As an example, we experimentally correlated drop heights with the onset of cavitation nucleation in different cell culture setups. We find that the critical drop height decreases by about 7 times when the height of cell culture media increases from 1 cm to 4 cm. This result indicates that lower drop heights induce the critical pressure (a material related property) with increasing cell media height and, as a result, implies that impact-induced pressure in biological systems depends on their size due to inertial effects. Another interesting experimental observation is that cells near cavitation bubbles uptake propidium iodide, typically not permeable to live cells, which we suspect is due to transient damage to the cell membrane. Our study shows that in addition to acceleration - which is the most commonly used criteria for blunt injury - size of biological systems, e.g., head size, should be appropriately considered for accurate assessment of potential injuries due to mechanical impact.

The present study delineates preparation, characterization, and application of calcium alginate (CA) - Gum tragacanth (GT) beads carrying phenolic compounds from B. alba plant for oral drug delivery. The CA- GT beads were prepared by ionic gelation method and its physicochemical characterization was carried out by SEM, EDAX and FTIR analysis. The swelling analysis and in vitro degradation assay revealed variation in the percentage of swelling and degradation rate respectively with the change in the GT concentrations in beads. Variation was prominent in compositions with higher GT concentrations.
A critical examination of the drug release profile revealed that the presence of GT in the formulation retarded the release rate of phenolic compounds. Extract loaded formulations were tested against osteosarcoma cells (MG-63). Cytotoxicity data, live dead staining, nuclear condensation-fragmentation, DNA fragmentation (by TUNEL assay) and cell migration study together confirmed the therapeutic potential of the CA-GT formulations. The above findings suggest that B. alba phenolic extract loaded CA- GT beads could be used as the natural therapeutic source for cancer treatment and an ideal source of nutraceuticals.

In general, the vitiligo is diagnosed by wood lamp test which is also known as ultraviolet (UV) light test. However, UV light in the wavelength under 400-nm is not visible to the human eyes and cameras. Thus, it is difficult to obtain accurate UV images for analyzing lesions of vitiligo patients. The light incident into the human skin has different penetration depth depending on the wavelength, which means the information of the reflected light differs according to the incident light wavelength. If there were any cameras or sensors that are capable of obtaining UV reflected images of the lesion, more deliberate information such as lesion size, depth, and shape could be obtained through a depth profile according to the penetration depth of human skin along the wavelength range.
Here, we developed a novel UV camera applying blue-light emitting quantum-dots (QD) to conventional Si CMOS image sensor (CIS). Synthesized zinc-blend-structure QD absorbs UV light below 400-nm wavelength and emits the 433-nm-wavelength blue-visible-light by energy-down-shift mechanism, having a quantum yield of 75.2%, full-width half maximum (FWHM) of 20.7 nm. The reflected light from the object passes through the QD filter before entering the photodiode of the Si CMOS image sensor. The QD filter completely passes the visible light and only absorbs UV light and QD emits the blue-visible-light. In other words, the QD filter converts UV light, which the conventional image sensors cannot detect, into the blue light so that the image sensor can detect it. This process presents that the image sensor receives a further increased blue pixel’s signal caused by UV intensity. The difference of blue pixels intensity between with and without QD filters can be considered as a detected UV intensity.
In this work, we diagnosed and treated lesions of 2 vitiligo patients through our new QD UV camera. We photographed the UV images of their hands and faces and analyzed them in cooperation with dermatologists. Based on the analysis, the patients were recommended for proper treatment and we tracked their treatment progress for 2 months. It was observed that the UV images of the vitiligo lesions showed lower UV pixel intensities than the intact normal skin, which means that the location, size and distribution of vitiligo lesions could be clearly identified through the UV image. The UV pixel intensities effectively showed information of epidermis and dermis where vitiligo lesions mainly exist. Melanocytes reflect and scatter ultraviolet rays to protect the skin from harmful UV rays. So, in UV images of normal skin, melanocytes reflect UV, resulting in high UV intensity. However, in the case of vitiligo lesions, there is less reflection of UV light due to the lack of melanocytes, resulting in low UV intensity. In addition, we can also estimate the extent of melanin pigment deficiency by this UV intensity analysis. The regions with relatively high UV pixel intensity and those with relatively low UV intensity were clearly distinguished in the vitiligo region. The relatively lower UV intensity means that the deficiency of melanocyte is relatively large, indicating that lesions are more severe. We confirmed that the novel QD UV camera can accurately discriminate the lesions of vitiligo from the intact region, and also can quantify the severity of the lesion. It is expected that the UV images taken by the QD UV camera will provide guidelines for the diagnosis and treatment of vitiligo. In addition, UV pixel analysis is also considered to be valid for several skin disorders that can be diagnosed by the UV light test such as tinea capitis, erythrasma, albinism, etc.
^*This work was financially supported by the Brain Korea 21 Plus Program in 2018.

Trauma brain injury (TBI) is a problem in public health locally and globally. The consequences of this pathology are serious and its mortality rate is high. TBI severity is classified using the Glasgow coma scale. This scale assigns points to the patient’s eye opening, motor and verbal response based on a qualitative test. Although this method is part of several TBI protocols, its application is susceptible to errors associated to the specific physiological response of each person. Misclassification of the initial TBI severity will in turn impact the therapeutic management causing in many cases delays in proper treatment. In this research work, we are proposing a quantitative method to classify the severity of the TBI, using highly specific serum biomarkers in order to reduce misclassification as well as to provide a fast, easy to use tool for monitoring the patient's progress. An electrochemical immunosensor have been developed by means of a specific antibody against S100B protein. S100B is a calcium binding protein present in the central nervous system; its concentration increases within the initial hours of the TBI event, its value as TBI biomarker have been demonstrated before. The biosensing platform was created by electrochemical polymerization of a nanodiamond enhanced polyaniline matrix, this matrix was then functionalized with antiS100B by crosslinking reaction after APTMS-GA (siloxane 3-aminopropyltriethoxysilane - glutaraldehyde) initial surface modification. The amount of antibody was optimized for the specific surface active area using different depositing dilutions, the results showed that after 5ugmL no statistically significant increment in the electrochemical signal is observed. Response of the biosensor to the antigen was recorded using impedance spectroscopy experiments in 0.2 M KCl and 10 mM potassium ferrocyanide as supporting electrolyte. Our results showed the characteristic semicircle behavior at the high frequency range associated to the redox probe of the electrolyte; also, the Warburg line has been observed at the low frequency range.

Functional brain changes represent a significant social and economic burden that has increased considerably over the last decades. The development of new scientific capabilities that are able to analyze brain structure and connectivity has recently revitalized the neuroscience field. Biomaterial based ex vivo models of healthy and diseased brain tissue are among those tools that may help study the complexity of cerebral tissue to better diagnose, prevent, and treat brain diseases (1). We have established engineered brain tumor biomaterials based on methacrylamide-functionalized gelatin hydrogels, and used microfluidic-forming techniques to generate platforms that combine transitions of biophysical and biomolecular properties found in the glioblastoma tumor microenvironment, from the core to the tumor margins (2). Moreover, this biomaterial approach is able to monitor the response of patient-derived xenograft (PDX) cell populations with different molecular signatures; in particular, we have analyzed the amplified and the constitutively activated vIII mutant EGF receptor. We have also characterized PDX response to targeted inhibitors, such as erlotinib, a tyrosine kinase inhibitor that specifically blocks EGFR.

The ex vivo biomaterial platforms that recreate transitions in the native GBM tumor microenvironment have shown the capacity to describe cell response to therapeutic inhibitors and classify tumors response to local gradients in extracellular matrix properties (3, 4). This in vitro tumor model is able to analyze the relationship between the molecular weight of matrix-bound hyaluronic acid (HA) and the invasive phenotype of GBM tumor cells. We found that cell growth, motility, and proteomic responses of GBM cells within our platform were significantly altered by HA molecular weight. These results provide additional insights regarding the importance of extracellular microenvironment in the invasive potential of glioblastoma tumors (5). Recently, we demonstrated that repeated dosing of erlotinib promotes expression of PDGFRb in EGFR vIII mutant cells and that extracellular HA plays a favorable role in the inhibition of EGFR, through STAT3 deactivation (6).

In addition to tumor tissue samples, these gelatin-based platforms permitted the survival and growth of the large nucleus gigantocellularis (NGC) neurons, considered as the 'master cells' for initiating CNS arousal. The better understanding of NGC neurons might help to understand particular characteristics of certain brain disorders (7). Until now, the lack of a physiologically relevant microenvironment for in vitro culture has been associated with unusual cell morphologies and cell death. We showed that NGC neurons were able to survive and differentiate in the presence of HA, and without the assistance of growth factors (8). In this context, 3D in vitro cultures of NGC neurons may allow the analysis of the extracellular matrix contributions and growth factors to their survival, growth and development. Due to the success of these platforms to recreate cerebral tissue peculiarities, we have also cultured neural stem cells within gelatin – HA hydrogels and we are currently working on the development of neurodegenerative diseases models.

References
(1) Pedron S and Harley BAC (2018) Editorial: Biomaterials for brain therapy and repair. Front. Mater. 5:67.
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(7) Inna Tabansky, Yupu Liang, Joel N. H. Stern, Donald W. Pfaff et al. (2018). PNAS, 115 (29) E6900-E6909.
(8) Magarinos AM, Pedron S, Pfaff DW, Harley B et al. Front. Mater., 28 June 2018

Electronic devices are one of the most promising candidates for chemical and biological sensor platforms because of its high sensitivity, and application to Internet of Things (IoT). However, a major challenge to developing such sensor systems is the use of battery power owing to its limited life-time, inconvenience of recharging, and insufficient battery level to operate the integrated sensor system.
In this study, we suggest self-powered biosensors by integration of n-IGZO/p-Si photodetectors and enzyme-based colorimetric reactions, which are operated by light sources in daily life environment such as fluorescent light and sunlight. A colorimetric reaction was performed at a polydimethylsiloxane vessel located at the upper side of the IGZO/Si photodetector, which is physically apart from the photodetectors. Photocurrent changes of the photodetectors are induced by the colorimetric reaction depending on target concentration, which enables quantitative analysis. The self-powered biosensors showed high sensitivity towards glucose level in real human samples without matrix effect due to physical separation of the photodetectors and colorimetric reaction part. This sensor platform could pave the way for highly sensitive portable IoT biosensors operating without batteries.

Recently the interest in bio-degradable Magnesium alloys as an alternative implant material has increased. Since no surgery is needed to remove the inserted implant, the risk of bacterial infection and the medical burden is decreased. In particular, anti-adhesion is one of the prerequisite properties in medical applications to avoid bacterial colonization on the implant. In this study, biocompatible Mg-Zn-Ca alloys are synthesized by squeeze casting and hydrophobic nano-SiO₂ particles are coated on the alloy surface. In the process, a porous SiO₂ layer is formed and its condition controlled via dip-coating. The human pathogen most frequently found on implant materials, Staphylococcus Aureus (S. Aureus), is prepared through the pour-plating method to evaluate the anti-adhesive properties and the bacteria on the surface are counted by scanning electron microscopy. The result shows that hydrophobically treated SiO₂ particle layers significantly mitigate the bacterial attachment compared to a bare Mg surface and a SiO₂ particle layer.

Introduction: Extracellular matrix (ECM) alterations accompany the onset and progression of many diseases. In this work, we sought to develop tissue engineering-based approaches to mimic the disease environment in order to perform prospective analyses that probe the mechanistic role of the ECM in disease etiology. Our specific focus was on calcific aortic valve disease (CAVD), whose pathogenesis is poorly understood, but is known to involve excessive glycosaminoglycan (GAG) accumulation early in the disease process. We implemented two different strategies to create tissue-engineered platforms that mimic elements of early CAVD: 1) construction of engineered environments using traditional “bottom-up” methods, which enable tuning and control of scaffold parameters but fail to capture the complexity of the native tissue. 2) construction of a top-down organ culture platform that retains native complexity, but lacks the ability to precisely control tissue composition.

Methods: To create scaffolds via the bottom-up approach, we copolymerized methacrylated gelatin with either methacrylated hyaluronic acid (HA) or chondroitin sulfate (CS) at concentrations intended to replicate the GAG enrichment found in early CAVD (1x GAG = healthy valve; 4x = diseased) and seeded them with valvular interstitial cells (VICs). To create top-down scaffolds, we transduced scaffolds to overexpress hyaluronic acid synthase-3, an enzyme responsible for HA synthesis. Porcine aortic valve leaflets were treated with feline immunodefficiency virus (FIV) containing a vector for either GFP (FIV.GFP) or human hyaluronic acid synthase-3 (FIV.HAS3) and then cultured in vitro for up to 7 days. Cultures were also treated with low density lipoprotein (LDL) or oxidized LDL (oxLDL), which mimic CAVD risk factors, and analyzed for overall ECM composition, compressive modulus, cell phenotype, and cytokine/growth factor production.

Results: Analysis of CS-enriched scaffolds demonstrated that pathological CS enrichment alone was not sufficient to promote a change in VIC phenotype or secretion of inflammatory factors. Instead, the main function of increased CS appeared to be sequestration of oxLDL. Sequestered oxLDL increased the secretion of multiple inflammatory cytokines (e.g., IL-6, IL-8, PDGF-BB, MCP-1) by VICs, thereby implicating oxLDL as a potent factor in regulating pathological VIC behavior. Moreover, a positive feedback loop was uncovered where oxLDL increased CS production, which further increased oxLDL retention. These findings suggest that increased CS in early CAVD plays a central role in regulating the onset of inflammation.

Meanwhile, a different role for HA enrichment was demonstrated in both bottom-up and top-down scaffolds. ECM-edited valve organ cultures were generated to achieve a 4-fold enrichment in HA content, with the level of HA enrichment regulated by the amount of virus administered. This result demonstrated the ability to tailor the ECM composition of intact, 3D tissues, thereby providing a novel disease-mimicking culture platform. In both culture approaches, HA enrichment served to stimulate vascular endothelial growth factor (VEGF) production by the VICs, but HA was not involved in LDL/oxLDL sequestration or subsequent inflammatory activity. Instead, HA-induced VEGF production was sufficient to stimulate the proliferation and tubulogenesis of endothelial progenitor cells, suggesting that increased HA in early CAVD may promote the pathological angiogenesis that is a hallmark of this disease.

Conclusions: We developed complementary tissue-engineered platforms to mimic disease pathogenesis, including a novel approach to use intact organ cultures as semi-tunable biomaterials. These platforms were applied to reveal new information about CAVD pathogenesis. The applications of these approaches extend beyond valvular disease, as there are many pathologies which involve selective enrichment of specific ECM components.

Molecularly targeted nanoparticles that produce X-ray excited optical luminescence, also known as radioluminescence, enable unique multimodal imaging capabilities not available with current clinical technologies. These functional nanomaterials, often comprised of lanthanide-doped nanophosphors, have also been evaluated as energy mediators for deep tissue photodynamic therapy (PDT) and other X-ray stimulated therapies. As both imaging probes and therapeutics, the application of radioluminescent nanoparticles (RLNP) is limited by their photophysical properties. Here we present the synthesis and characterization of lanthanide-doped nanoparticles with core-shell structure, incorporation of high-Z elements, and their surface modification with biocompatible polymers to improve functional properties for biological applications. In this radioluminescent platform, optical emission can be tuned by selection of dopants, such as Eu³⁺ and Tb³⁺. Utilizing a thermal degradation of organometallic precursors, uniform cubic (~11 nm) nanocyrstals were obtained and observed by transmission electron microscopy (TEM). To evaluate the radioluminescence of these nanophosphors a suspension of the particles was irradiated at 130 kV while an emission spectrum was obtained by a spectrophotometer. Emission was improved significantly through protection from surface quenching of luminescent centers. In addition, surface modification with a layer-by-layer approach was developed and cell specific uptake was evaluated using folic acid as a model targeting agent. Based on these results, we discuss the feasibility of X-ray luminescence computed tomography (XLCT) and X-ray excited PDT.

Cancer-associated fibroblasts (CAFs) are one of the major types of stromal cells within the tumor microenvironment that support tumor progression by secreting pro-tumorigenic cytokines.^1-2 In addition, CAFs also play a key role in stromal remodeling during tumor progression, including increased deposition and crosslinking of collagen.³ In general, the matrix mechanics have been shown to affect various cellular behaviors.⁴ However, to our knowledge, limited studies have been carried out to investigate the effects of matrix biomechanical changes on CAFs behavior, particularly in three-dimensional (3D) environment as well as the downstream paracrine crosstalk with the surrounding tumor cells remains elusive. Hence, in this study, a collagen/alginate interpenetrating network (CoAl-IPN) hydrogel system was employed to investigate the phenotypic and functional characteristics of CAFs under 3D environment setting. In general, collagen and alginate have different crosslinking mechanisms, which will allow the precise control of the hydrogel biomechanical properties without affecting its biochemical properties.

Results showed that hydrogel with Young’s Modulus from ~108 to 898 Pa was successfully fabricated by varying the crosslinking density of alginate which preserving the biochemical properties. Subsequently, CAFs were encapsulated within the CoAl-IPN hydrogel system. Interestingly, a highly polarized CAFs morphology was observed within compliant CoAl-IPN hydrogel (i.e. uncrosslinked system), but not rigid hydrogel system (i.e. highly crosslinked). This is in contrast to 2D cell culture findings, in which stiffer substrate will promote the cell spreading. In addition, the compliant CoAl-IPN hydrogel system also affected the CAFs behavior at molecular level with enhanced nuclear translocation of the mechano-sensitive YAP/TAZ protein and expression, as well as the up-regulation of other CAFs classical markers (i.e. α-SMA and FAP).

Our findings also demonstrated that CAFs cultured in this compliant ECM was able to promote the spreading of MDA-MB-231 tumoroids by its paracrine activity. Collectively, these findings show that the CAFs phenotype and functional characteristics are highly plastic, which depends on the biomechanics of the microenvironment. Moreover, these understanding from the tumor stroma mechanic-bioactivity relationship studies can shed the lights on the restraint the pro-tumorigenic effects of CAFs in cancer therapy.

1. Quail, D. F.; Joyce, J. A., Microenvironmental regulation of tumor progression and metastasis. Nat Med 2013, 19 (11), 1423-37.
2. Mbeunkui, F.; Johann, D. J., Jr., Cancer and the tumor microenvironment: a review of an essential relationship. Cancer Chemother Pharmacol 2009, 63 (4), 571-82.
3. Malik, R.; Lelkes, P. I.; Cukierman, E., Biomechanical and biochemical remodeling of stromal extracellular matrix in cancer. Trends Biotechnol 2015, 33 (4), 230-6.
4. Calvo, F.; Ege, N.; Grande-Garcia, A.; Hooper, S.; Jenkins, R. P.; Chaudhry, S. I.; Harrington, K.; Williamson, P.; Moeendarbary, E.; Charras, G.; Sahai, E., Mechanotransduction and YAP-dependent matrix remodelling is required for the generation and maintenance of cancer-associated fibroblasts. Nat Cell Biol 2013, 15 (6), 637-46.

Three-dimensional (3D) porous scaffolds with large specific surface areas have received considerable interest due to their capabilities of effective cell attachment and growth. For their utilization, the cells are cultured over the pre-fabricated scaffolds, which typically not only accompanies a long-time process, but also arises the challenge to precisely position target cells inside the scaffolds. Here, we utilize an additive manufacturing technique to fabricate nanofibrous 3D porous scaffolds in which target biological species are located. The fabricated platform shows the selective permeability such that biological species are confined inside, while chemical species can pass through the porous layers, due to the much smaller nanofibers than biological species. As such, the platform performs excellent biochemical conversion under the repeated processes. Moreover, the fabricated scaffolds are electrically conductive, which enables cell monitoring by using electrochemical measurement techniques. Our strategy can provide a guideline to improve cell utilization in many applications with biochemical conversion.

Acknowledgement
This work was supported by the Intelligent Synthetic Biology Center of Global Frontier Project of the National Research Foundation of Korea (NRF-2012M3A6A8054889), funded by the Ministry of Science and ICT of the Republic of Korea. This research was also supported by “Human Resources Program in Energy Technology” of the Korea Institute of Energy Technology Evaluation and Planning (KETEP), granted financial resource from the Ministry of Trade, Industry & Energy, Republic of Korea. (No. 20184010201710).

Oxidative stress during sepsis pathogenesis remains the most-important factor creating imbalance and dysregulation in immune-cell function, usually observed following initial infection. Hydrogen peroxide (H₂O₂), a potentially toxic reactive oxygen species (ROS), is excessively produced by pro-inflammatory immune cells during the initial phases of sepsis and plays a dominant role in regulating the pathways associated with systemic inflammatory immune activation. In the present study, we constructed a peroxide scavenger mannosylated polymeric albumin manganese dioxide (mSPAM) nanoassembly to catalyze the decomposition of H₂O₂ responsible for the hyper-activation of pro-inflammatory immune cells. In a detailed manner, we investigated the role of mSPAM nanoassembly in modulating the expression and secretion of pro-inflammatory markers elevated in bacterial lipopolysaccharide (LPS)-mediated endotoxemia during sepsis. Through a facile one-step solution-phase approach, hydrophilic bovine serum albumin reduced manganese dioxide (BM) nanoparticles were synthesized and subsequently self-assembled with cationic mannosylated disulfide cross-linked polyethylenimine (mSP) to formulate mSPAM nanoassembly. In particular, we observed that the highly stable mSPAM nanoassembly suppressed HIF1α expression by scavenging H₂O₂ in LPS-induced macrophage cells. Initial investigation revealed that a significant reduction of free radicals by the treatment of mSPAM nanoassembly has reduced the infiltration of neutrophils and other leukocytes in a local endotoxemia animal model. Furthermore, therapeutic studies in a systemic endotoxemia model demonstrated that mSPAM treatment reduced TNF-α and IL-6 inflammatory cytokines in serum, in turn circumventing organ damage done by the inflammatory macrophages. Interestingly, we also observed that the reduction of these inflammatory cytokines by mSPAM nanoassembly further prevented IBA-1 immuno-positive microglial cell activation in the brain and consequently improved the cognitive function of the animals. Altogether, the administration of mSPAM nanoassembly scavenged H₂O₂and suppressed HIF1α expression in LPS-stimulated macrophages and thereby inhibited the progression of local and systemic inflammation as well as neuroinflammation in an LPS-induced endotoxemia model. This mSPAM nanoassembly system could serve as a potent anti-inflammatory agent, and we further anticipate its successful application in treating various inflammation-related diseases.

Multifunctional nanoparticles integrating cancer cell imaging and treatment modalities into a single platform are recognized as a promising approach; however, their development currently remains a challenge. In this study, we synthesized magnetic field-inducible drug-eluting nanoparticles (MIDENs) by embedding superparamagnetic iron oxide nanoparticles (Fe₃O₄; SPIONs) and cancer therapeutic drugs (doxorubicin; DOX) in a temperature-responsive poly (lactic-co-glycolic acid) (PLGA) nanomatrix. Application of an external alternating magnetic field (AMF) generated heat above 42 °C and subsequent transition of the PLGA polymer matrix (T_g = 42–45 °C) from the glassy to the rubbery state, facilitating the controlled release of the loaded DOX, ultimately allowing for simultaneous hyperthermia and local heat-triggered chemotherapy for efficient dual cancer treatment. The average size of the synthesized MIDENs was 172.1 ± 3.20 nm in diameter. In vitro studies showed that the MIDENs were cytocompatible and especially effective in destroying CT26 colon cancer cells with AMF application. In vivo studies revealed that the MIDENs enabled enhanced T₂contrast magnetic resonance imaging and a significant suppression of malignant tumor growth under an AMF. Our multifunctional MIDENs, composed of biocompatible substances and therapeutic/imaging modalities, will be greatly beneficial for cancer image-guided thermo-chemotherapy applications.

Piezoelectric materials have the ability to convert mechanical energy to an electrical signal leading to their use in a variety of microscale sensor systems having biological and medical applications such as pressure sensors, chemical sensors, and biosensors. Additionally, the ability of piezoelectric materials to convert electrical signals to mechanical energy has led to their use as microactuators. Microcantilever sensors have been increasingly used in biomedical microelectromechanical systems (bioMEMS) to measure small scale mechanical movement. One application is in the determination of contractile force of various muscle tissues using in vitro human-on-a-chip systems, which have typically used imaging methods or optical laser deflection of cantilevers for calculating contractility. Incorporation of piezoelectric materials into cantilever-based mechanical sensors would enable a direct electrical readout during muscle contraction. Moreover, these cantilevers could be used in reverse, as actuators, to impose a stress on tissues attached to the device. This work describes the design and fabrication of a piezoelectric bioMEMS cantilever which can act as either a sensing element for tissue contraction or as an actuator to apply mechanical force to cells. A piezoelectric sensing element composed of a thin film of aluminum nitride (AlN) was fabricated on a microscale silicon cantilever which was subsequently integrated with human primary skeletal muscle or human iPSC-derived cardiac muscle to monitor the contractility of the muscle tissues. An electrical signal generated by the piezoelectric element from the contracting muscle (skeletal or cardiac) was interfaced to a commercial amplifier for recordings. Force generation was calculated from cantilever deflection data generated by spontaneous or stimulated contraction of the cells. The ability of the piezoelectric cantilevers to serve as actuators for applying mechanical stress to tissue constructs was evaluated using an optical beam displacement method to measure the deflection of the piezoelectric cantilever under a periodic waveform of applied voltage. This novel bioMEMS sensor has the potential for incorporation into human-on-a-chip systems for basic physiological investigation and pharmaceutical compound development, cosmetics toxicity testing, and personalized medicine.

We describe the evaluation of lactose modified graphene oxide (GO-AL) as potential drug carrier targeted to hepatic asialoglycoprotein receptor (ASGPR). Structural-modification and functional analysis of GO-AL were performed. The structure and morphology of the composite were analyzed by scanning electron microscopy (SEM), and atomic force microscopy (AFM), while Raman spectroscopy was used to track the chemical modification. For the safe application of GO-AL an evaluation of the cytotoxic effect and specific interaction of glycoconjugate were also studied. The SEM and AFM analysis of the GO showed graphene sheets with 2-3 nm layer size and few of them reached the 4 nm. Raman spectrum showed the characteristic peaks of graphene oxide at 1608 cm^-1 and 1350 cm^-1 corresponding to G and D bands, respectively. Null cytotoxic effects were observed for GO samples proving the biocompatibility. The biorecognition of GO-AL and the specific absorption to HepG2 cells were confirmed. The high efficacy of conjugation, hemolytic safety, and specific recognition described here to lactose modified GO represents a potential drug delivery vehicle to hepatic tissue.

Cancer is one of the leading causes of death around the world. The lifestyle and external factors can increase the cancer risk, no matters economical status and develop of countries. Current therapies such as radiation and chemotherapy alone or in combination are still unsatisfactory, and survival of cancer patients is still low. Therefore, it is necessary to develop a novel therapy that improves survival rates and the life-quality for cancer patients. In this sense, smart nanocarriers constitute an evolving possibility for controlled drug delivery to the tumor, which minimizes side effects in the patient. Among its properties, these systems have the option to respond reversibly to external factors, such as temperature, pH, ionic strength, and others. The chitosan-graft-poly(N-vinylcaprolactam) (Cs-g-PVCL) is a water-soluble copolymer that could comply with requirements for biomedical applications. Our aim in this work was to develop stimuli-responsive nano-formulations from Cs-g-PVCL copolymer as a potential drug carrier and evaluate the influence of the preparation conditions on their thermosensitive properties. The Cs-g-PVCL nanoparticles (Cs-PVCL-NP) were prepared by ionotropic gelation using TPP as a crosslinking agent. The Cs-PVCL-NP were characterized by dynamic light scattering (DLS), ζ-potential, atomic force microscopy, IR-FT spectroscopy, and thermogravimetric analysis. The results showed that morphological characteristics of Cs-PVCL-NP are dependent of the concentration of Cs-g-PVCL and the copolymer:TPP volume ratio. The thermosensitive properties of the nanoparticles evaluated by turbidimetric method and DLS showed a phase-transition at temperatures close to physiological conditions. Additionally, the nanomaterial showed the capability to encapsulate and release doxorubicin. Accordingly, this nanomaterial offers interesting properties for the development of smart chemo-drug controlled delivery carrier, which is a potential alternative method to treat cancer.

Dielectric breakdown is a physical phenomenon observed when applied voltage exceeds dielectric strength of insulator. In case of insulating films, it is known that nanopores can be formed caused by dielectric breakdown, which can be used widely for biosensor applications. One of the techniques to form nanopores is using dielectric breakdown of insulating films, which can be simple in comparison with other methods. In this study, silicon nitride (Si₃N₄) with thicknesses of 10 and 20 nm and hexagonal boron nitride (h-BN) with thicknesses in the range of 10 to 40nm were used for nanopores formation. In order to observe the dielectric breakdown, an insulating film was inserted between two completely isolated chambers, and potassium chloride (KCl) solution was filled in the chamber. After that, we could observe dielectric breakdown of the insulators after elapsed time when a specific voltage is applied to insulating films. Some molecules can pass through nanopores and generate unique signals as they go through the pores. As a result, we could observe the translocation of DNA through nanopores formed by dielectric breakdown and this can be applied to the development of nanopore-based biotechnologies.

Recently, in the medical area, alternative antibacterial solutions have been investigated, because the excessive use of antibiotics has caused the microorganisms to have a greater resistance to these antibiotics. In this research work the Ag nanoparticles effect is monitored as a function of the incubation time by using different concentrations of colloidal solutions on the bacterium Pseudomonas aeruginosa (ATCC-27853) in order to identify the minimum inhibitory concentration. The nanoparticles were synthesized using Pelargonium spp extract as a reducing agent and nanoparticle concentrations in colloidal solution of 1: 2, 1: 4, 1: 8 and 1:16 were evaluated, in order to verify the effect on pseudomonas aeruginosa by means of the count of Colony Forming Units (CFU) during 0, 2 and 4 hours of incubation of the bacteria. Average diameters of nanoparticles <50 nm were obtained, which were characterized with an Electronic Transmission Microscope JEOL JEM-1010. The tests showed that the 1: 2 concentration ratio showed a 100% inhibition at the time of mixing the colloidal solution with the growing bacteria and during the four hours of monitoring. Whereas in the 1: 4 ratio, 100% inhibition was observed up to 2 hours after the nanoparticles were mixed with the bacteria. For the rest of the dilutions, there were gradual inhibitions as a function of time, however they were not 100%. Finally, the determination of the effect of the Ag nanoparticles evaluated in this work suggest carrying out cytoxicity assays in cellular biological systems for applications of biomedical use.

Intense exploration of cellulose nanocrystals (CNCs) to assemble biomimetic natural chiral nematic materials is conducted for fabrication of flexible biophotonic materials, but achieving both high structural robustness and controlled optical iridescence is still challenging. Short CNCs facilitate uniform helicoidal nanostructure for the optical enhancement but lower loading transfer efficiency and poor mechanical performance, while long CNCs benefit the mechanical performance but sacrifice iridescence because of physical entanglement. Herein, we report a composite CNC materials, which is capable of self-assembling into uniform iridescent helicoidal CNCs films with superior mechanical performance. Such hierarchical composition enable composite CNCs films to possess highly ordered helicoidal architecture for brilliant iridescence, combined with an unusually high mechanical strength, high elastic modulus, and toughness, more than an order of magnitude higher than those reported early for purely CNC films with preserved iridescent. This study reveals an efficient strategy to overcome the conflicting trends in mechanical and optical performance of traditional chiral CNC films and their conversion to robust chiral photonic nanocomposites without introduction of external synthetic polymer components as supporting matrices.

In this study, a novel approach to block a carboxybetaine (CB)-based zwitterionic segment in polyurethanes (PUs) to address biocompatibility, antifouling properties and tunable mechanical properties has been developed. This zwitterionic PUs were synthesized via the polycondensition of diisocyanate with CB-based diol and CB-based triol, respectively. The hydrolysis of both diol and triol-segments at the interface generates zwitterionic CB functional groups that provide excellent antifouling properties via the enhanced hydration capacities of CB groups. Thermogravimetric analysis and differential scanning calorimetry measurement show the high thermal stability of poly(carboxybetaine urethane)s with up to 305 ^oC degradation temperature. Tunable mechanical properties and water uptake were corresponding to the feed ratio of CB-based diols and triols. This approach demonstrates a new idea to block CB functional groups into polyurethane without complicated synthetic route or post-modification. It also provides a better understanding on relationship between structure and function of zwitterionic polyurethanes. Owing to its biocompatibility, thermal stability, tunable mechanical properties and great antifouling properties, this CB-based polyurethane can be emerged as a promising candidate for biomedical application. It also can be an alternative replacement for those PUs without antifouling properties.

The development of nanoparticles which can be used as a stimuli responsive drug carrier for the treatment of different diseases has been an emerging area of research interest. In our study, we have designed a chitosan-bilirubin micelle (nanoBil) carrying losartan for the treatment of liver fibrosis which is responsive to intrinsic reactive oxygen species (ROS). Bilirubin is hydrophobic in nature and its carboxyl group was conjugated to the chitosan amine group using the EDC-NHS chemistry to form amphiphilic conjugate nanoBil. Losartan has proved to be an angiotensin I receptor blocker having hepatic fibrosis reduction capability and it is being used as the therapeutic payload in this study to form nanoBil-losartan micelle. The ROS responsive Nano micelle can release loaded drug inside at the liver fibrotic area, hence liver fibrosis is characterizerd with high ROS levels. The release characteristic of nanoBil-losartan was tested by ROS generator to confirm losartan release. The reduction in the hepatic stellate cell activation after the treatment of nanoBil-losartan were analyzed based on the alpha smooth muscle actin (α-sma) expression both in vitro (LX2 cell line) and in vivo studies. Hydroxyproline level as well as histopathological evaluation of the liver showed a decreased collagen content in the treated groups. Both the intravenous and intraperitoneal administration of the nanoBil-losartan resulted in a decreased level of fibrosis in thioacetamide (TAA)/Ethanol induced liver fibrosis C3HeN mice model. The in vivo and in vitro results suggest that the ROS stimuli responsive nanoBil nanoparticle could be a potent therapeutic option for inflammatory diseases like liver fibrosis which are abundant of ROS.

Accuracy and precision are essential requirements for chemical analysis. Herein, we developed highly precise amperometric sensor for point-of-care diagnosis of blood glucose. Stainless steel 304 needle was employed as a sensing electrode since it possessed decent electrical properties with low material cost. Especially, we found that it displayed extremely low non-faradaic background current in electrochemical condition as well as highly precise and sensitive current response specifically toward hydrogen peroxide. In blood condition, we confirmed the stainless steel 304 needle immobilized with glucose oxidase showed current signal at -0.15 V (vs SCE) upon the addition of glucose. The amperometic current signal from the modified electrode was highly specific to the blood glucose with negligible response towards interfering species. Moreover, the signal from the glucose was extremely precise with linear correlation in the diabetic blood sugar concentration range of 1-20 mM (y = 2.52328 x 10^-8x, R² = 0.9952) with negligible standard deviation (σ = 1.78 x 10^-9 A, n = 5, CV = 0.10 – 0.84 %). These results demonstrate that stainless steel 304 needle electrode hold great potential in electroanalytical biosensors based on redox enzymes relevant to hydrogen peroxide.

Abstract: As a new and hot spot, the nucleic acid nanotechnology plays a more and more important role in cancer targeted therapy because of its perfect biocompatibility, high sensitivity and functional richness. Including directional drug carrier and release, fluorescent tracking and so on. DNA condensation is a good way to get these kind of carriers. Metal complexes, such as Co(NH₃)₆³⁺, can induce DNA condensation. A recent study found that pyrophosphate magnesium, the RCA reaction by-product, can condense the long single stranded DNA into the tight micro structure. Based on this finding, we report a condensation of single-strand DNA with excessive magnesium ions and form the nanoparticles whose diameter is around 100nm. Furthermore, combined with the programmability of DNA sequence and structure, we finally get an functional Mg-DNA nanoparticle with high doxorubicin loading rate, high tumor-targeting property, high biocompatibility and low toxicity. Due to the wealth of DNA sequence design, this kind of DNA nanoparticles will have a great potential on nano-medical treatment in the future.
Key words: DNA condensation, tumor-targeted chemotherapy, nanostructure, drug delivery
Reference:
[1] Huang, Y.; Xu, W.; Liu, G.; Tian, L. A Pure DNA Hydrogel with Stable Catalytic Ability Produced by One-Step Rolling Circle Amplification. Chem. Commun. 2017, 53, 3038−3041.
[2] Guan, Y. L.; Rui, L. G.; Liang, N. J.; Hui, C. DNA Condensation Induced by Metal Complexes. Coordination Chemistry Reviews., 2014,281,100-113.
[3] Hu, R.; Zhang, X. B.; Zhao, Z. L.; Zhu, G. Z.; Chen, T.; Fu, T.; Tan, W. H. DNA Nanoflowers for Multiplexed Cellular Imaging and Traceable Targeted Drug Delivery. Angew. Chem., Int. Ed. 2014, 53,5821−5826.

Nanocomposites comprising of biopolymers and calcium phosphate cements (CaP) are promising due to their biocompatibility, non-toxicity, biodegradability and suitable mechanical properties for biomedical applications. In here, a new composite material was synthesized with carboxymethylcellulose (CMC) and gelatin (GEL) as the liquid phase and CaP based powder as the solid phase. In this study, the effect of the addition of different wt% of GEL including 0, 5, 10, 20 in the liquid phase was investigated on the physical properties of the nanocomposites. Physico-chemical characteristics of materials were determined by using Fourier transform infrared (FTIR) spectroscopy, scanning electron microscopy (SEM) and mechanical tests. Swelling analysis was performed after 1, 8, 16, 24 and 48 hours and the degradation of samples was studied after 7 and 14 days. FTIR results showed that there was a physical interaction between CMC and GEL with H bonding which was evident from the peak at 3288cm^-1. Disruption in GEL structure was observed from the band at 1600-1400 cm^-1 Disruption of GEL structure may increase the interaction between CMC and GEL molecules. After mixing of solid and liquid phases, negative charges on CMC and Ca²⁺ molecules start to interact with each other and produce attraction sites for PO4^3- molecules. This lead to an accumulation of hydroxyapatite-like structures. A homogenous and porous microstructure was observed by using SEM in all samples. Mechanical tests showed that GEL improved the strength of the samples up to 20 wt% of GEL. The addition of 20 wt% of GEL decreased the mechanical properties. The compressive strength values were found up to approximately 6 MPa. Swelling results revealed that increasing in GEL concentration cause decrease in swelling until the 16^th hour after that 10wt% GEL samples had the lowest swelling which was approximately 28%. Finally, degradation studies indicated that the highest degradation rate was for 10% GEL incorporated samples. Addition of further GEL also reduced the degradation rate. Overall, the addition of GEL improved physical properties of the samples for potential biomedical applications.

The PEGylated OCN (PEG-OCN) was well dispersed in water as nanodots with a lateral dimension showed strong photoluminescence in the visible region. Recently, thin 2D materials, such as graphene derivatives, have become an emerging nanosized platform in various biological applications, such as biosensors and their unique fluorescent, electronic, and morphological features, and high surface areas.
In particular, graphitic carbon nitride (g-C₃N₄), composed of triazine or tri-s-triazine building blocks containing alternating sp²-hybrid carbon and nitrogen atoms, is a stable metal-free 3D layered material. However, this g-C₃N₄ has a less water dispersibility because stable 2D tri-s-triazine network. So we first make oxidized carbon nitride(OCN) material by using KMnO₄ in the presence of H₂SO₄. Thus 3D OCN has a oxygen abundant structure.
Fluorescent materials are being used for the optical/fluorescence imaging of living cells and animal models. As such, the development of heavy-metal-free, water dispersible, and biocompatible imaging probes is still important. Carbon nitride (C₃N₄) is used as a bioimaging probe due to its suitable optical properties, thus enhancing its biocompatibility and dispersibility in aqueous media is required.
In this study, we incorporated short-chain polyethylene glycol (PEG) groups onto a carbon nitride network by the simple N-alkylation of hexaethylene glycolic mesylate with nucleophilic nitrogen atoms on oxidized carbon nitride . Cell-viability testing confirmed that these “heavy-metal-free” organic nanodots were highly biocompatible and noncytotoxic. In particular, the developed nanodots could provide clear confocal images of RAW 264.7 cells without weakening cell activity and displaying any aggregation in a range of concentrations (25–100 mg/mL ) with bright-green emission in the cytoplasm.

Microbial carbohydrates have a variety of characteristics and original functionalities comparing with carbohydrates produced by animals and plants. Microbial carbohydrates are natural, non-toxic, biodegradable and biocompatible polymers, GRAS(Generally Regarded As Safe) and their structural diversities lead to a variety of functions. Recently, many novel applications have been developed using microbial polysaccharides such as drug delivery systems, hydrogels, nanoparticles, and materials for tablet-pressing process in the pharmaceutical and biomedical industries as well as in the bio-nano engineering. Microbial polysaccharides can be utilized as stabilizer, emulsifier, thickener, gelling agent in food industries. The Microbial Carbohydrate Resource Bank (MCRB) was established to investigate and collect various functional polysaccharides and microorganisms in order to widely utilize the microbial resources. MCRB also take a role as resource bank through microbial modification and offering a storage facility for various microbial carbohydrates and microorganisms. In addition, MCRB will contribute the advancement of industrial fields using carbohydrates and provide microbial resources into various researchers to encourage basic and applied researches.

Improving the mechanical properties of calcium phosphate cement (CPC)-based injectable bone substitutes (IBS) is crucial to expand their application in the treatment of bone defect. In this study, the effect of graphene oxide (GO) on the mechanical properties of CPC-based injectable bone substitutes (IBS) were studied by using X-ray diffraction (XRD), scanning electron microscopy (SEM), dynamic mechanical analysis (DMA), rheological measurement techniques. Also, the produced IBS samples were analyzed by means of setting time and temperature, injectability, bioactivity and biocompatibility studies. To this end, IBS samples were prepared with 0.5, 1.0, 1.5 and 2.0 wt% GO incorporation. According to the results of the DMA analysis, incorporation of GO increased the compressive modulus of IBS samples after setting of the cement phase. For example, while the compressive modulus of 0.5wt% GO incorporated IBS samples was ∼0.2 MPa, the compressive modulus of 2wt% GO incorporated IBS samples was ∼0.7 MPa. A sharp increase of the storage modulus (G’) and viscosity (Pa.s) in rheological measurement was observed which indicates the gelation temperature and gelation time. The gelation temperature and time were decreased effectively with the addition of GO. XRD analysis of GO incorporated IBS samples showed that the peak intensity and the grain size of CPC increased with respect to wt% of GO incorporation. SEM analysis showed that GO added IBS samples had a denser microstructure. Moreover, alamar blue assay showed that all the prepared samples were biocompatible. Although the increase of wt% of GO reduced the injectability, overall; the addition of GO as a reinforcement has a positive effect on the mechanical properties and handling of injectable bone substitutes.

Nanotechnology-based biosensors offer the ability to leverage a vast numbers of sensing elements per unit area in a single sensor device offering the potential develop new sensing methodologies. When coupled with a smart readout mechanism they offer the ability to mimic sensitive olfactory systems that relies on an interplay between sensitive receptors and synaptic processing of olfactory information. Surface enhanced Raman scattering (SERS) - vibrational spectroscopic sensing - can simultaneously sense large numbers of small molecules, which can be differentiated based on their vibrational signature; thus it doesn’t require capture agents such as antibodies and aptamers. Yet, SERS spectra are complex and difficult to attribute to individual molecules in a complex mixture.
To establish a new sensing paradigm using SERS, we first will present how 2-dimensional physically activated chemical (2PAC) assembly uses electrohydrodynamic flow as a long range driving force to enable controlled molecular cross-linking between nanospheres and thereby overcomes long standing challenges in reproducible nanomanufacturing of plasmonic ‘nanoantennas’ serving as ultrasensitive receptors on SERS chips. 2PAC achieves uniform (<10% relative standard deviation) billion fold enhancements - capable of measuring a single molecule of interest – in elements spanning mm². Large (>10,000 spectra) datasets can thus be acquired from the 2PAC fabricated SERS sensors that are critical for reliable training data for machine-learning. We show how this offers the ability to rapidly and reliably process the rich spectral data. For example, when applying deep convolutional neural network (CNN) analysis of SERS spectra, we are able to quantify (not simply detect!) rhodamine 800 down to 100 femtomolar concentration. We further demonstrate that transfer learning can be used to generalize a rhodamine 800 trained CNN to quantify concentrations of methylene blue.
We will then show that this SERS + machine learning approach enables early detection of biofilm formation on contaminated surfaces. For example, cultures from the opportunistic pathogen Pseudomonas aeruginosa are flowed over SERS sensors in a microfluidic channel. Machine learning analysis of SERS spectra are able to detect biofilm formation as early as 3 hours after inoculation. We do so by quantifying the concentration of pyocyanin, a metabolite produced by P. aeruginosa, that we show is correlated with the accumulation of biomass. Surface-attached bacteria exposed to a bactericidal antibiotic were differentially less susceptible after 10 h of growth, indicating that these devices are useful for early intervention of bacterial infection. Further, we present that the complex vibrational signature of metabolites after antibiotic exposure provides the ability to distinguish between antibiotic resistant and susceptible strains. While new antimicrobial strategies are being developed to combat antibiotic resistance, here we present a promising parallel strategy, using signals of dysregulation of core metabolic functions as evidence of antimicrobial efficacy. Consider that previously Raman spectroscopy has outperformed mass spectroscopy and DNA sequencing in discriminating between Acinetobacter baumannii strains; aspects of the rich spectral data are needed to capture non-genomic data for differentiation. We will present results showing that the complex vibrational signature of metabolites can be used to distinguish antibiotic resistant P. aeruginosa from antibiotic susceptible P. aeruginosa. This is achieved after just 30 minutes of antibiotic exposure at the minimum inhibitory concentrations using linear discriminant analysis of SERS spectra from bacterial lysate.

In this study, the effect of addition of 0, 0.5, 1, and 2 weight (wt) % of carboxylated multi-walled carbon nanotube (f-MWCNT) was investigated on the physical properties of carboxymethyl cellulose (CMC)/calcium phosphate (CaP)-based cements for biomedical applications, particularly in tissue engineering. The structural and mechanical properties were characterized by using techniques including Fourier Transform Infrared Spectroscopy (FTIR), Scanning Electron Microscopy (SEM), mechanical tests, and swelling-degradation tests. According to FTIR results, all samples had peaks at 3394 cm ^-1 and 1085 cm^-1 which confirmed hydrogen bonding between CMC and CaP-based cement and well crystallization of CaP, respectively. A homogenous and uniform morphology of samples was observed from SEM images, as well as a highly porous structure. Moreover, the addition of f-MWCNT improved the mechanical properties and the compressive strength of f-MWCNT incorporated nanocomposites reached approximately up to 9 MPa. Swelling test was performed after 1, 8, 16, 24, and 48 hours. The nanocomposites prepared with the addition of 1 wt % f-MWCNT had the lowest % swelling, which was found as 32.40 % after 48 hours. On days 7 and 14, the degradation studies indicated that the degradation rate was slowed down with the increase of wt % of f-MWCNT in the nanocomposites. Our results show that the prepared nanocomposites may be potential candidates for a range of tissue engineering applications, including bone regeneration.

Zinc Oxide (ZnO) nanocomposites have attracted much attention for various biomedical applications, such as drug delivery, bio-sensor, diagnosis and therapy due to biocompatibility and multifunctionality. [1] Therefore, various synthesis methods of this promising material have been extensively studied to control nanostructures. [2] However, there are limitations to synthesize nanostructures by combining ZnO with other polymeric materials, especially fiber structured polymer. Herein, we introduce a unique method for synthesizing ZnO nanowires (NWs) on polylactic acid (PLA) microfiber and applied them for cancer therapy.
The Hierarchical structured nanocomposite composed of ZnO NWs and PLA microfibers are synthesized by two simple and effective methods. First, randomly oriented PLA microfibers containing ZnO nanoparticles seeds are electrospun on collector. Subsequently, ZnO NWs are directly grown from the seeds embedded in fiber via hydrothermal method. In addition, the NWs are stably fixed on the fibers and their densities are also successfully controlled according to the amount of seeds. Since the process of growing ZnO NWs from the seeds inside the fiber is a quite unique phenomenon, their growth mechanism and kinetics are also addressed in this paper.
Moreover, this inorganic-organic nanocomposite provides a new approach to immunotherapy because there biodegradable, biocompatible and multifunctional properties. Conventionally, in therapy application, ZnO-based composites are directly injected into target cells to release embedded drugs or operate therapy using cytotoxicity. However, this nanocomposite plays a role in transferring only antigens on the nanowires to the cells. We demonstrated the nanocomposites induce immune response about specific target biomarkers and inhibit tumor growth in vivo. ZnO and PLA nanocomposite reduced immune suppressive regulatory T cells and improved the infiltration of T cells into tumor tissues when compared to mice immunized with PLA fibers coated with the antigen. This novel structured nanocomposite provides a new possibility in immunotherapy

Reference
[1] P. Sharma et al., Nanoscale, 9, 15371 (2017)
[2] Z. L. Wang, J. Phys.: Condens. Matter 16, R829 (2004)

Soft gels that mimic properties of biological samples are increasingly utilized as tissue simulants for the assessment of potential damage, e.g., blunt injuries, to a human body against rapid mechanical inputs. As a result, capabilities to accurately characterize and predict transient, dynamic response of soft materials under fast loading rate conditions have become critical for such biomedical applications. Here we present (1) a new experimental method for characterizing dynamic cavitation nucleation in biologically relevant soft materials; (2) quantitative characterization on the critical acceleration that corresponds to the onset of cavitation nucleation; (3) pressure gradient and transient bubble dynamics due to mechanical impact. Utilizing a drop tower based system, we have quantified the critical acceleration in soft gels while concurrently visualizing material deformation during cavitation events. Our experiments show that the critical acceleration increases with increasing concentration of gels. Note that gel stiffness increases with increasing concentration. For stiffer gels, more energy is needed for local deformation of a gel associated with bubble expansion and, as a result, larger mechanical inputs, i.e., higher accelerations, is expected to trigger cavitation nucleation. Also, we have empirically showed that pressure gradient in the gel is proportional to the square of the depth of the sample. Our study on how biological soft materials respond to rapid mechanical inputs is an important step to understanding possible underlying injury mechanism(s) in blunt trauma and cavitation-induced brain injury.

The recent advances in point-of-care diagnostics have led to the development of high-performance electrical sensors that can provide fast measurements with low cost and multiplexing capabilities. However, detection in complex environments is still challenging due to the non-specific adsorption of proteins that may foul the sensor surface leading to signal attenuation. Therefore, nucleic acid purification from biological samples is often a crucial, yet a laborious initial step. In general, a simple and reliable approach for base mismatch detection is yet to be achieved. To address these challenges, we present an approach for a biosensor that consists of two stages for mismatch detection in complex environment with simple purification process occurs in the first stage then the second stage investigates at a molecular level the target’s sequence using single molecule break junction (SMBJ) technique. The first stage involves a nanostructured electrode, where the electrode morphology has been demonstrated to be biofouling resilient. This coating enables both detection of specific targets in a complex biological sample and their subsequent purification using electrochemical methods. The morphology of the electrodes allows small molecules to pass and participate in the bio-electrochemical reaction happening at the electrode surface. Nanoporous gold (np-Au) electrodes can be modified with a DNA capture probe which enables sensitive detection and capture of complementary DNA or RNA targets in the presence of complex media (blood). The surface-bound DNA:DNA or RNA:DNA can then be released by electrochemically cleaving the thiol-gold linkage, eluting the hybrids from nanoporous matrix for further analysis. Complementary to the electrochemical screening approach, single molecule conductance measurements can provide a molecular insight that is unavailable using electrochemical methods. The single-molecule break junction (SMBJ) system employs a movable, nanostructured electrode tip which makes direct contact to a gold substrate. In these measurements, charge transports between the electrodes through a single bridged-molecule. The magnitude of the current passing through the molecule will change drastically compared to tunneling gap. Here, the eluted sample from the first stage can be investigated using SMBJ which can differentiate between perfectly matched vs mutated targets (3-base mismatch). Our results show the difference in conductance values between perfect and mismatch targets in complex media. The advantages of combining these two approaches offer the ability of detection of single-nucleotide polymorphisms, simultaneous identification of multiple targets, high signal-to-noise ratio and low limits of detection.

Injuries involving the peripheral nervous system (i.e. nerve lesions or transection) are estimated to count for 2.8% of all trauma cases in the United States; when considered alongside neurodegenerative disorders such as Parkinson’s disease, nerve damage has affected no fewer than 20 million people in the US alone. At present, the “gold standard” clinical intervention is a nerve autograft, which addresses the worst morbidities of nerve injury but is not without drawbacks. Autografts are limited in their ability to repair complex injuries, they often introduce new sensory deficits or neuropathy in the donor region, and may present problems with structural mismatch between donor and repaired nerve fascicles. Artificial nerve conduits are of great interest as a replacement or supplement to autograft-based techniques, but functional recovery remains an issue with current conduit designs. Incorporation of biocompatible Conducting Polymers (CPs) has been proposed to improve neurite guidance and improve conduit performance due to CPs’ electrical, physical, and chemical properties. The roughness of CP surfaces may have an important influence on the rate and directionality of axonal regeneration, but few works have specifically addressed this potential effect.

We have investigated two common biocompatible CPs, poly(pyrrole) (PPy) and poly(3,4-ethylenedioxythiophene) (PEDOT), fabricated using galvanostatic (GSTAT) and potentiostatic (PSTAT) methods, and characterized the surface roughness of these two polymers. While PPy is well-known for its electrical conductivity and physical stability, PEDOT displays superior chemical stability and electrical conductivity, making it an ideal candidate for artificial conduits.

Electrodeposition was performed on microfabricated gold electrodes using a solution of EDOT or Py monomer and poly(styrenesulfonate) under GSTAT (0.1 to 1.5 mA/cm²) or PSTAT (0.5 to 0.95 V) deposition modes over intervals ranging from 1 to 10 minutes. Each surface was analyzed with materials confocal microscopy (MCM) to measure the CP film thickness and surface roughness. Additionally, the impedance and charge storage capacity (CSC) of the films were measured with impedance spectroscopy and cyclic voltammetry, respectively. Our results suggest a dual dependence of all measured properties (roughness, thickness, impedance, and CSC) on both deposition time and applied charge. For example PPy/GSTAT films on Michigan neural electrodes, roughness (Rq) increased from 2.14±0.25nm to 12.0±0.84nm; CSC also increased from 5.7mC/cm² to 98.7mC/cm²; finally, impedance at 1000Hz decreased from 1.84e5 Ω to 3.83e4 [MRA1] [KAM2] Ω. These measurements were taken from films deposited at 1min, 0.1mA/cm² and 10min, 1.5mA/cm² respectively. PPy properties exhibited generally linear trends, and PEDOT properties tended to display nonlinear trends. Dorsal root ganglion (DRG) cells will be cultured on these CP films and neurite outgrowth will then be assessed as a function of surface roughness. These results can be leveraged to improve the control of regenerating axons via surface cues.

The use of nanocomposite materials can contribute significantly to improve the quality of population life. In this context, nylon 6,6 (N6) is one of the most important engineering plastics in the automobile industry and has been investigated as a bone tissue scaffold. Trisodium trimetaphosphate (STMP) can be used as potential candidates for dental applications due to the anticaries action of STMP when present in a biocompatible scaffold as N6. Therefore, the insertion effect of different STMP concentrations in a N6 polymeric matrix was evaluated and correlated to the physicochemical properties of nanocomposites. STMP nanoparticles were prepared by mechanical milling for 48h. N6 and its nanocomposites were prepared by electrospinning technique, while, the N6/STMP nanocomposites were processed by adding 2.5, 5 and 10% w/w (STMP:N6). The milling processing reduced the particle size of the STMP powders without affecting its crystalline structure. Particle size was reduced of micrometric to nanometric scale after mechanical milling producing particles with ~70 nm and spherical morphology. The phase structure was analyzed by FTIR technique and nuclear magnetic ressonance spectroscopies employing ¹³C-CPTOSS (cross polarization with total sideband suppression). ¹³C NMR all chemical shifts were well resolved and were assigned according to N6. By FTIR spectra it was observed characteristics peaks of N6 and STMP, showing that, STMP was incorporated in N6. The morphology it was analyzed by Scanning Electron Microscopy (SEM) technique. SEM images showed the formation of nanofibers in N6 and its nanocomposites with ~150 nm of thickness for N6 and thickness higher for N6-STMP nanocomposite, showing the presence of STMP homogeneously distributed over the nanofibers. The thermal behavior was analyzed by TGA and DSC technique. Thermogravimetry analyses demonstrated improved thermal stability of N6-STMP nanocomposites with higher STMP concentration according to its barrier effect. Additionally, the increase in the glass transition temperature in N6-STMP nanocomposites indicated the reduction of polymeric chains mobility. The mechanical properties were evaluated, and N6-STMP -2.5% nanocomposite presented higher elastic modulus, elongation at rupture and tensile strength, presenting as a potential candidate in dentistry. These findings showed a new approach to add STMP nanoparticles by electrospinning in a polymeric matrix, forming stable nanofibers with potential application in dental biomaterials. With this methodology, it was possible with this methodology to insert the STMP nanoparticles in a polymeric matrix and increase the physicochemical properties of the nanocomposites formed.

Gold nanoparticles have been studied extensively for the strong capability in cancer treatment and their potential in the anti-bacterial field.^1-2 Due to the ease of synthesis, chemical stability, and controllable toxicity, gold nanomaterials, especially branched gold nanoparticles, have attracted enormous interest as new biomaterials.^3-4 This research focuses on the controlled synthesis of branched gold nanoparticles and their antibacterial applications.
The surfactantless synthesis method reported by Xie and Wang was utilized to produce the 10 nm level branched gold nanoparticles.⁵ The 20 nm to 60 nm level branched gold nanoparticles were prepared by seed-mediated synthetic route.⁶ To investigate the anti-bacterial effects, the branch number and length were modulated by the pH value and 4-(2-hydroxyethyl)-1- piperazineethanesulfonic acid (HEPES) buffer concentration. Then, the branched gold nanoparticles were characterized by Ultraviolet–visible spectroscopy (UV-Vis) and transmission electron microscopy (TEM). Moreover, the 10 nm to 60 nm level spherical gold nanoparticles were produced by the method reported by Brown and Natan and utilized as the control group.^6-7 Furthermore, branched nanoparticles with diverse sizes have been utilized to reduce bacterial growth (e.g., Staphylococcus aureus, Staphylococcus epidermidis, methicillin-resistant Staphylococcus aureus, and Streptococcus) for the anti-bacterial applications.

Reference
1. Penders, J.; Stolzoff, M.; Hickey, D. J.; Andersson, M.; Webster, T. J., Shape- dependent antibacterial effects of non-cytotoxic gold nanoparticles. Int J Nanomedicine 2017, 12, 2457-2468.
2. Qian, X.; Peng, X. H.; Ansari, D. O.; Yin-Goen, Q.; Chen, G. Z.; Shin, D. M.; Yang, L.; Young, A. N.; Wang, M. D.; Nie, S., In vivo tumor targeting and spectroscopic detection with surface-enhanced Raman nanoparticle tags. Nat Biotechnol 2008, 26 (1), 83-90.
3. Alkilany, A. M.; Murphy, C. J., Toxicity and cellular uptake of gold nanoparticles: what we have learned so far? J Nanopart Res 2010, 12 (7), 2313-2333.
4. Chithrani, B. D.; Ghazani, A. A.; Chan, W. C. W., Determining the Size and Shape Dependence of Gold Nanoparticle Uptake into Mammalian Cells. NANO LETTERS 2006, 6 (4), 7.
5. Xie, J.; Lee, J. Y.; Wang, D. I. C., Seedless, Surfactantless, High-Yield Synthesis of Branched Gold Nanocrystals in HEPES Buffer Solution. Chem. Mater 2007, 19, 7.
6. Maiorano, G.; Rizzello, L.; Malvindi, M. A.; Shankar, S. S.; Martiradonna, L.; Falqui, A.; Cingolani, R.; Pompa, P. P., Monodispersed and size-controlled multibranched gold nanoparticles with nanoscale tuning of surface morphology. Nanoscale 2011, 3 (5), 2227-32.
7. Brown, K. R.; Natan, M. J., Hydroxylamine Seeding of Colloidal Au Nanoparticles in Solution and on Surfaces. Langmuir 1998, 1998 (14), 3.

TiO₂ has been widely used as a photosensitizer for solar energy, hydrogen generation, removal of pollutants, and in biomedical applications such as photodynamic therapy (PDT). Despite its successful application in PDT, TiO₂ does not distinguish between healthy and cancer cells, hence killing both. Modification by a targeting cancer agent may provide a more effective solution. In this work, hollow TiO₂ nanospheres (HTiO₂NS) are obtained by using a carbon template-mediated method and functionalized with Zinc phthalocyanine (ZnPc) and folic acid (FA) for enhancing its photocatalytic activity and increasing selectivity of cancer cells, respectively. To study the attachment of FA and ZnPc onto TiO₂NS Fourier-transform infrared (FTIR) spectroscopy is used to characterize the functionalization of HTiO₂NS. The crystal structure was analyzed using X-ray diffraction (XRD). Size distribution, morphology, and hollowness are characterized by transmission electron microscopy (TEM). Brunauer–Emmett–Teller (BET) was taken with N₂ to measure the surface area and study surface porosity of produced HTiO₂NS. Furthermore, functionalized HTiO₂NS with ZnPc-FA are tested in vitro under UV illumination to measure the reactive oxygen species (ROS) generation with a fluorescent probe. A biological test was performed via a microculture tetrazolium assay (MTT) to measure cell death and asses the effectiveness of functionalized HTiO₂NS by ZnPc-FA. The results show that HTiO₂NS modified by ZnPc-FA can serve as a potential platform for targeted photodynamic therapy.

We fabricated microchannels with overhang and helical geometries by sequential sacrificial molding of two substrates printed by a fused deposition modelling (FDM) 3D printer. Using a 3D model consisting of two complementary materials, we fabricated a sacrificial mold with free-hanging geometries removable in water after the removal of the other substrate used as a support material. 3D printing has become an indispensable tool to fabricate microfluidic devices. In particular, FDM 3D printing has been used to create entire microfluidic devices and to fabricate sacrificial molds in removable materials. Although 3D printing may confer wide freedom of design, the geometry of the attainable sacrificial mold is still restricted to the capabilities of the 3D printer and associated materials. For example, sacrificial molds made with a single material in FDM cannot offer microchannels with overhang features. 3D geometries such as dual helixes remain challenging to print using FDM printing, and requires the use of auxiliary tools to facilitate printing of geometry suspended in air.

To address these limitations, we demonstrated a method that we termed dual sacrificial molding. In dual sacrificial molding, we used two complementary materials (high impact polystyrene (HIPS) and polyvinyl alcohol (PVA)) to fabricate 3D microchannels with free hanging geometries. PVA was used to model the microchannel, and HIPS was used as a support material for the PVA mold. HIPS in the sacrificial mold was firstly removed by soaking in limonene to reveal the PVA mold harboring the free-hanging geometries. We showed that PVA was intact and retained the original design after the exposure to limonene. Thereafter, the PVA sacrificial mold was embedded in PDMS and subsequently removed by soaking in water. Using this method, we fabricated two representative designs of microchannels such as dual pyramidal networks of microchannels and dual helical microchannels, both of which were not achievable by the 3D printing of sacrificial molds based on a single material.

The capabilities of dual sacrificial molding to produce microchannels harboring free hanging geometries relies on the complementary pairing of two sacrificial materials and respective solvents used to remove them. Unlike direct printing of a sacrificial mold with a single material, our method extends the freedom of design of microchannels attainable by FDM 3D printing. We highlight the potential of using ubiquitously available FDM 3D printers and commercially available filament as sacrificial materials to fabricate complex 3D microchannels in a range of matrices relevant to chemical and biological applications. More specifically, the complementary pairing of PVA and HIPS allowed the retention of a water-soluble mold which is biocompatible and versatile to use with a wide plethora of matrix materials. To this end, we envision that the method presented here could serve as an essential method to fabricate complex vasculature in biologically relevant matrices which would be difficult to achieve with a single sacrificial material and other methods of 3D printing.

Given that dysregulation of mechanics contributes to diseases ranging from cancer metastasis to lung disease, it is important to develop methods for measuring the molecular forces applied by cells. This is a difficult problem, as molecular forces are at the piconewton scale (1 kcal/mol ~7pN*nm). To address this issue, we have developed a suite of molecular tension probes that are engineered using PEG polymers, oligonucleotides, and proteins. Tension probes employ plasmonic nanoparticles, which provide significant improvement in sensitivity. Fluorescence polarization spectroscopy and super-resolution imaging provide the highest resolution maps of cell forces. I will describe the application of these probes in the study of integrin and T cell receptor mechanobiology. Finally, I will also demonstrate applications of these materials in the area of personalized mechano-pharmacology.

Engineering cell culture environments that mimic both the 3D and the dynamic nature of tissues is a major challenge in tissue engineering. Even with advanced manufacturing techniques such as 3D printing, it is difficult to produce 3D structures of soft hydrogels, which are otherwise ideal for cell culture.

Self-folding materials allow the generation of cell-friendly structures with 3D complexity across a range of length scales that are difficult to achieve with printing and lithography. Folding is induced by differential stresses generated within a flexible or elastic material. Polymeric materials, and hydrogels in particular, are well-suited for self-folding applications because they undergo pronounced volume and modulus changes in response to cell-friendly stimuli such as water, pH or temperature, to generate differential stresses within. While self-folding hydrogels have been reported, heterogeneities that lead to differential swelling must be pre-patterned into the material, and thus the shape change occurs as soon as the material is places in a physiologically relevant environment (lacks temporal control). Furthermore, only a few examples utilizing these materials with live cells have been reported.

We have designed photodegradable hydrogels incorporating ortho-nitrobenzyl moieties that permit encapsulation or cell seeding on swollen planar films. Thin sheets are programmed to undergo a 2D to 3D shape change in response to light which induces irreversible folding of the hydrogel sheet due to differential swelling of the degraded and non-degraded regions. This bottom-up approach enables the use of simple 2D precursors to create biologically relevant 3D structures such as tubes and folded containers for cell encapsulation. Unlike other shape-changing polymer systems, such as poly(N-isopropylacrylamide), the o-NB hydrogel system requires no laminated bilayers, pH change or elevated temperatures to induce shape-change. Furthermore, photodegradation based shape-change enables external temporal control of folding that can be induced at time points of choice during cell culture.

Biological imaging is an indispensable component of modern medicine which can shed light even on the molecular level on the biological phenome and can be utilized for the early diagnosis of a wide gamut of deadly diseases such as cancer. Conventional imaging methods such as X-ray radiography, computed tomography (CT), and magnetic resonance are commonly used in the clinic to diagnose physiological and anatomical abnormalities. Despite their high spatial resolutions, these methods are incompetent compared with optical imaging methods as the latter offer high sensitivity, real-time screening, and intraoperative feedback while also being safe because of the lack of harmful radiation. Of importance is optical imaging in the first biological window where the interference from tissue scattering and tissue autofluorescence is minimal, leading to the enhanced depth of penetration.
In particular, the great potentials of fluorescent imaging have led to the development of organic dyes and quantum dots (QDs) with high quantum yield. However, small molecules tend to get renally cleared in a short time span making the imaging practically cumbersome. On the other hand, although QDs might address the fast-renal exertion of the small organic dyes, their toxicity due to the presence of heavy elements has always been a bottleneck slowing their development for the in vivo imaging.
Herein, we disclose for the first time the synthesis of ultrasmall nanoparticles coined as “oligodots”. The term was adopted after comprehensive mass spectroscopy analysis revealed the oligomeric nature of these nanoparticles. Their hydrothermal synthesis with slight modification in the solvent, resulted in photoluminescence tuning of the derived oligdots with green- (G-Dot) and red- (R-Dot) emitting fluorescent properties. Size exclusion chromatography, AFM and HRTEM all corroborated the formation of 1.5±0.5 nm NPs. 2D Excitation- emission contours revealed that R-Dots have an when excited at 530 nm while the G-Dots have . Detailed fluorescence spectroscopic methods such as time resolved photoluminescence spectroscopy (TRPL) and fluorescent correlation microscopy were performed to fully characterize their photophysical properties. The incubation of the NPs with mammalian breast cancer cell (MCF-7) and fluorescent confocal imaging indicated the high performance of these nanaoparticles for in vitro cell imaging.
Having demonstrated their feasibility for biological studies, we carried out an in vivo study in nude mice. The G-Dot and R-Dot were injected intradermally in each of the mouse’s flanks and were imaged on IVIS instrument. Spectral unmixing was utilized where the correct localization of the oligodots was achieved indicating their multiplexing potential. Moreover, the R-Dots were injected in the tail vein and the mice were imaged at various timepoints up to 24 h (. The results confirmed the feasibility of the in vivo imaging and the proper time window clearance over 3 h. Histopathological examination of the major harvested organs, revealed no sign of cytotoxicity of any kind. Considering the desirable spectroscopic properties, safety and proper pharmacokinetics, these nanoparticles can offer a new platforms as novel fluorescent imaging agents on multiple levels.

Defects of the cranial skeleton occur frequently secondary to trauma, stroke, cancer, and congenital anomalies. Such defects result in significant neurological, psychological, social, and vocational burdens necessitating reconstruction. Current clinical options for reconstruction include autologous bone or alloplastic materials. However, the former is limited by tissue availability and donor site morbidity while the latter is plagued with frequent complications including extrusion of material, infection, and failure particularly in hostile wounds such as those following radiation. The limitations of both options provide the impetus for investigation in calvarial regeneration. Despite decades of research, contemporary regenerative strategies consisting of expanded stem cells and growth factor cocktails delivered by scaffolding materials have not attained clinical translation due to lack of efficacy compared to autologous bone, high cost, excessive time consumption for harvest and ex vivo progenitor cell expansion, and the untoward effects of supraphysiologic dosages of growth factors. With the increasing knowledge of the instructive capabilities of the extracellular matrix, our laboratory is primarily interested in identification of regenerative biomaterials, characterization of mechanistic interactions between materials and biology, and clinical translation of regenerative biomaterials tailored for craniomaxillofacial defects. Our laboratory recently demonstrated the ability of an extracellular matrix-inspired material composed of nanoparticulate mineralized collagen glycosaminoglycan (MC-GAG) for regeneration of massive calvarial defects in the absence of ex vivo progenitor cell expansion or exogenous growth factor supplementation. We further showed that the mechanistic basis for MC-GAG induced osteogenic differentiation was dependent on an autogenous activation of the bone morphogenetic protein receptor (BMPR) signaling pathway via expression of a panel of BMP ligands by the osteoprogenitors themselves. Our current research interests involve: 1. Inorganic ion requirements and signaling in osteogenic differentiation induced by MC-GAG 2. Negative regulatory pathways for osteoclastogenesis and osteoclast activation induced in progenitor cells undergoing osteogenic differentiation on MC-GAG 3. Development of Good Laboratory Practices-compliant protocol for testing the safety and performance of MC-GAG in animal cranial defect models.

Over the past decades, the increase in kidney stone (KS) formers has raised the importance to understand the biomineralization process responsible for urolithiasis^[1]. Calcium oxalate (CaOx) crystallization – KS main inorganic compound^[2] – has largely been characterized under batch synthesis conditions that cannot be regarded as biomimetic with respect to the microscale environment in the kidney and to the urinary flow. A chemical model considering these features would provide, in fine, the biomedical community with a predictive platform for suitable and reliable diagnosis.
In this work, we used a reversible microchannel to mimic the collecting duct in the nephron where CaOx stones can form due to supersaturated levels in calcium and oxalate ions^[3]. Within the channel, CaOx crystallization was induced under co-laminar mixing of Ca²⁺_(aq) and Ox^2-_(aq) ions matching pathological concentrations – i.e. hypercalciuria and moderate hyperoxaluria. Scanning electron microscopy and Raman spectroscopy were used to support our investigations. They showed that CaOx crystals precipitate in a mixture of monohydrated whewellite (CaC₂O₄.H₂O, COM) and dihydrated weddellite (CaC₂O₄.2H₂O, COD) in the microchannel, similar to what is observed by the physicians^[4]. In situ information on the kinetics of CaOx crystal growth could be acquired in our microfluidic system. They confirmed the effect of the hydrodynamic conditions as well as of the presence of chelating molecules on the growth kinetics and the final chemistry (phase, shape) of the formed CaOx crystals. Finally, in a trial to achieve a more complex biomimetic model (formation of KS on a Randall’s plaque), hydroxyapatite was grown in the microchannel and the CaOx crystal formation was investigated.

[1] V. Romero, H. Akpinar and D. G. Assimos, Reviews in Urology 2010, 12, e86-e96.
[2] M. Daudon, Annales d'urologie 2005, 39, 209-231.
[3] M. Daudon, P. Jungers and O. Traxer, Lithiase urinaire, Lavoisier, Médecine Sciences publications, 2012, p.
[4] M. Daudon, E. Letavernier, V. Frochot, J.-P. Haymann, D. Bazin and P. Jungers, Comptes Rendus Chimie 2016, 19, 1504-1513.

Although cancer nanomedicine has been developed for decades, translation has been slow. Delivery efficiency is one of the major obstacles. A recent study analyzing the cancer nanomedicine systems in the past 10 years reveals that only 0.7 % of administrated nanoparticles could reach the tumor tissue; under this condition, the amount of nanoparticle required for treating a cancer patient would be impractical for cancer therapeutics. As a result, an approach to significantly enhance the delivery efficiency is an unmet need, particularly for certain intractable cancers, such as glioblastoma (GBM), which only has limited options for treatment and the outcome remains unsatisfactory. To tackle this issue, we report an active delivery approach using the tumor-homing capability of mesenchymal stem cell (MSC) to boost the delivery efficiency as well as the retention of nanomedicine system to GBM.

MSC is capable of sensing the chemotaxis generated by cancer cells because of its upregulation of the specific receptors, such as C-X-C chemokine receptor type 4 (CXCR4). By forming a MSC spheroid via the microfluidics-based double emulsion method, we found that MSC’s CXCR4 expression was enhanced by 6 times, thereby giving MSC spheroid 2-fold faster migration toward the GBM cells. In addition, this allowed us to easily incorporate the nanoparticles with MSC during the spheroid formation within the confined double emulsions. We designed a DNA-templated micelle, formed through the calcium phosphate nanoprecipitation of PEGylated DNA templates, to carry the chemotherapeutic drug, mitoxantrone (MTX). The DNA template enabled efficient loading of MTX through intercalation, and the PEG component confined the precipitation to improve micelle’s stability. Incorporation of DNA-templated micelle did not compromise MSC spheroid’s migration capability, and more than half of the loaded micelles were stable in the extracellular matrix part of the spheroid, which could be carried by the MSC spheroids during migration toward the GBM cells.

We further engineered the MSC to secrete the cytotoxic protein, tumor necrosis factor-related apoptosis-inducing ligand (TRAIL). Co-delivery of TRAIL and MTX using the hybrid spheroid killed the GBM cells efficiently and specifically in vitro. In an in vivo GBM model, this hybrid spheroid approach gave nearly 100-fold improvement on micelle retention. At three-week post-administration, 11 ± 3.2 % of the micelles remained in the tumor site when the mice treated with our MSC/micelle hybrid spheroids, while only 0.16 ± 0.03 % of the micelles was found in the tumor site when treated with the single MSC/micelle approach. Accordingly, the GBM tumor grew significantly slower when treated with the hybrid spheroids, reaching only 14% of the tumor size found in the control group at day 21, the end of the experiment. Collectively, our MSC/nanomedicine hybrid spheroid approach integrates the merits of both MSC- and nanoparticle- based therapy, enabling enhanced drug delivery to achieve a superior therapeutic efficacy in vivo.

To create the tunable interface for cell engineering, this study is focusing on how to control the surface morphology of PEM with limited conditions. First, the surface density of thiol self-assembled monolayer can be the first candidate to control the morphology of PEMs. Due to unconstrained self-assembly reaction, thiol group (such as MUA) first adsorb and cover the whole surface with lying down phase that undergoes a transition to the line-up configuration to form the complete monolayer. The contact angle shows the angle was decreased after 6 hours SAM because that lying down phase of the carboxylic acid directly on the gold surface and then gradually increasing after 24 hours and 48 hours SAM to form line-up phase. Furthermore, quartz crystal microbalance (QCM) show that different SAM density will affect the formation of the PEMs and lead to different physical characterizations from dissipation diagram. More compact SAM layers, the surface weight will be higher than the low-density film as well as the surface softness. To investigate the surface morphology, Atomic force microscopy (AFM) images showed dramatically different during 6 hrs SAM for a flat surface, 24 hrs SAM to form island surface, and 48 hrs SAM for highly compact wrinkle morphology. Besides, the ionic strength of buffer can be other factors to affect more detail condition of PEMs morphology. From QCM results, salt effect will reduce or increase the electrostatic interaction between poly(acrylic acid) and poly(lysine) that reduce or enhance the surface weight of PEM. Interestingly, the morphology of PEM can also be affected to form the dots, fibers, and wrinkle structures corresponding to the salt concentration. By controlling the density of SAM and ionic strength of the buffer, the surface morphology of PEM can be fine-tuning and utilized as the new interface for cell engineering. Fibroblast cell can be utilized as the first demonstration of this platform. By monitoring morphology, proliferation and differentiation of cells, we can know more details how the PEMs affect the cell behaviors.

Limitations with existing methods and materials used for nerve repair necessitate the design and characterization of novel materials that can enhance the desired neural responses, and, promote controlled and precise adhesion, growth, and directionality of neurons on artificial “smart” substrates. Recent published studies have demonstrated the impact of aerogel topography on PC-12 cells. On average, the neurite length of PC-12 cells grown on aerogels is longer than those cultured on control substrates such as tissue culture plastic dishes. Here, through experimental studies our team has identified and characterized a technique for patterning of collagen-coated mesoporous aerogel biomaterials through plasmonic photo-patterning. By combining the precision of plasmonic photopatterning with liquid crystal-coated aerogels, we have created well-defined patterns on silica aerogels. The technique utilizes plasmonic photomasks for surface photoalignment. Results describe the methodology related to creating collagen-liquid crystal gel mixture, photo patterning, and cell responses to predefined pathways created on aerogel susbtrates.

Advances in materials science, biomolecule delivery, and cell biology has enabled significant innovations within the field of tissue engineering and regenerative medicine over the past few decades. Nonetheless, minimal translation of tissue engineering-based therapeutics to the clinic has occurred. A significant challenge within tissue engineering is the difficulty in regenerating complex tissues with a heterogeneous structure and multiple cell types. To address this challenge, my research group is developing innovative polymeric biomaterials that can mimic the complex microenvironment of musculoskeletal tissues, specifically interfacial tissues (e.g. tendon to bone interface). I will discuss our recent efforts in the following areas: 1) using magnetic fields to spatially control electrospun fiber alignment in order to create materials with gradients in fiber alignment; 2) using photoconjugation techniques to spatially control biomolecule presentation and create materials with gradients in fiber surface chemistry; and 3) combining these techniques for independent spatial control over chemical and physical signals towards improved regeneration of musculoskeletal interfacial tissues.

Exosomes are small (few tens ~ hundreds nm) vesicles secreted by various cells in human body. Recent advances of nanotechnology unveiled that exosomes contain important biomarkers including miRNAs and different types of proteins specific to parental cells. Hence, increased number of studies are reported the potential of exosomes as a stable and reliable biomarker for various cancers.

Previous approaches are mainly focused on utilizing heterogeneous and complex exosomes derived from human cells / body fluids. Because of the innate complexity of exosomes samples, translational research is limited and these limiting factors can be categorized into 1. Isolation, 2. Detection and 3. Heterogeneity problems. For example, gold standard isolation method requires time-consuming ultracentrifugation yet its effects on exosome morphologies are not well-understood. Additionally, there are limited number of studies on differentiating cancer specific exosomes from other exosomes derived from normal cells.

In this talk, we will talk about our strategy of designing artificial exosomes through microfluidics mixer design for understanding exosome sensing with field effect transistor (FET) based biosensor device. Specifically, artificial exosomes with 40, 70, and 100 nm size with narrow size distribution (PDI = 0.01) are successfully synthesized with desired density of membrane proteins and membrane charge densities. We will discuss how exosomes’ size, charge density, and concentration of membrane proteins are correlated to sensing signals in the FET biosensor through combined small angle x-ray scattering and cryogenic transmission electron microscope techniques. Finally, we will discuss how these relations can be used to interpret sensing signals from urinary exosomes.

Bacterial surface contamination is a major cause of hospital-associated infections (HAIs), and antibacterial coatings can play an important role in reducing bacteria transmission via inanimate surfaces in healthcare settings. Coatings with nanosilver are deemed to have great potential as antibacterial surface coatings for many applications such as medical devices and food packaging. Antibacterial nanosilver coatings can be easily fabricated using polydopamine (PDA) as an anchor layer to reduce and electrolessly metalize silver ions into silver nanoparticles. We have prepared nanocopper using PDA as an anchor layer, and tested its antibacterial efficacy against the nanosilver coating. In simulated contamination of inanimate surfaces by droplets or touch as expected in a hospital environment, the nanocopper coatings exhibited a much higher antibacterial efficacy than those based on nanosilver. We further investigated the relative advantages of nanocopper-PDA coatings compared to Cu(II) ion-complexed PDA coatings in terms of efficacy against the common HAI pathogens, S. aureus and P. aeruginosa, durability as well as potential cytotoxicity. Such coatings of varying copper content were formed on poly(vinyl fluoride), latex as well as stainless steel substrates. When contaminated with droplets of the bacterial suspension, the nanocopper coating exhibited significantly higher efficacy (no viable bacteria observed after 30 min) than the Cu(II) ion-complexed PDA coatings of similar Cu content (~90% reduction in viable bacteria) due to a higher amount of Cu released by the former. However, the higher amount of Cu released by the nanocopper coating can give rise to cytotoxic effects. When contaminated with a bacteria-loaded dry surface, both surfaces were highly effective in killing the surface bacteria (no viable bacteria observed within 20-30 min). With sufficiently high copper loading (>4 μg/cm²), these Cu-containing coatings were able to maintain their antibacterial efficacy after 100 wipes with DI water. The fabrication of the Cu-containing PDA coatings can be readily scaled up for practical applications. Depending on the intended application, the Cu content as well as its physical form (ions or nanoparticle) can be selected to achieve an optimal balance between antibacterial efficacy and cytotoxicity.

Cardiovascular disease is the leading cause of death in the United States, yet biomaterials developed to address specific pathologies of cardiovascular diseases such as coronary artery bypass grafts and heart valves still suffer from blood compatibility issues, requiring replacement or life-time anti-coagulation therapy.¹ Previously, we have demonstrated that coating electrospun polyurethane vascular grafts with bioactive PEG-based hydrogel coatings confers both thromboresistance and promotes endothelial cell attachment.² However, our previous fabrication technique involved a molding process that presented significant challenges for industrial scale-up as well as difficulty in achieving thin, conformable coatings with sufficient homogeneity. In this study, we present a new method to assemble thin hydrogel layers on electrospun meshes through redox-initiated crosslinking mediated by the reaction between iron gluconate (IG) and ammonium persulfate (APS). We investigated IG adsorption and desorption from electrospun meshes, the effect of time on hydrogel coating thickness, and the conformability of this methodology to different geometries.
Iron Gluconate Desorption: Bionate was electrospun onto a rotating mandrel to produce highly aligned fibers ~1 µm thick. The resultant polyurethane meshes were cut to 5x10 mm sheets, held in custom 3D-printed clamps, and soaked in 1, 3, or 5 wt% IG for 15 minutes. Meshes were then dipped in methanol and dried with compressed air. IG release from meshes immersed in water for 2 minutes in 10 second intervals was monitored with UV-Vis spectroscopy (n=18 for each concentration).
Redox Hydrogel Coatings of Tunable Thickness on Electrospun Meshes: IG-coated meshes were immersed in a solution of 10 wt% poly(ethylene glycol) diacrylate (PEGDA, 3.4kDa or 6kDa) and 0.14% APS for times ranging from 10-120 seconds. After immersion, hydrogel coated meshes were immediately immersed in water to remove excess polymer and initiator. Hydrogel thickness was measured by sectioning composites and visualizing cross sections with a stereoscope (n=12 for each condition).
Conformable Coatings: To demonstrate the application of this approach for conformable coatings, tubular electrospun meshes (4 cm long, 4 mm diameter) were coated in IG and immersed in PEGDA+APS as described above. Thickness of the coatings was analyzed with a stereoscope (n=6).
IG release from meshes after 2 minutes desorbing was determined to increase from 0.06 ± 0.02 mg for a 1% IG soak to 0.46 ± 0.09 mg for a 5wt% IG soak. Hydrogel coating thickness increased from 70 ± 33 µm after a 10 second immersion in PEGDA 3.4kDa to 310 ± 66 µm after a 120 second immersion. Higher molecular weight PEGDA 6kDa coatings were 280 ± 27 µm thick after 60 seconds as compared to 180 ± 22 µm PEGDA 3.4kDa coatings at the same time. Hydrogel coatings were formed conformably on the inside of electrospun tubular electrospun vascular grafts with a thickness of 370 ± 62 µm.
In these studies, we demonstrate a tunable and scalable technique for fabricating thin and conformable hydrogel coatings on polyurethane meshes. We believe this technique can be extended to coat a range of materials with different chemistries and geometries. Coating cardiovascular devices with PEGDA hydrogel layers holds the potential to make these devices thromboresistant and able to adhere endothelial cells. Future work will include coating artificial heart valves and individual fibers as well as assembling tough hydrogel coatings with this technique.
References:
1. Benjamin, E. J.; et al. Heart disease and stroke statistics—2018 update: a report from the American Heart Association. Circulation 2018, 137 (12), e67-e492.
2. Browning, M. B.; et al. Endothelial cell response to chemical, biological, and physical cues in bioactive hydrogels. Tissue Engineering Part A 2014, 20 (23-24), 3130-3141.

Cells are mechanically stimulated by the surrounding environment of extracellular matrices, neighboring cells and varieties of possible external physical stresses, and respond dynamically toward such mechanical stresses change of biochemical signaling pathways induced with physical stimulation has named as “Mechanotrasduction”, and has a crucial effect on cell proliferation, invasion, migration, apoptosis and differentiation. Especially, cells on various artificial mechanical cues (different stiffness substrate, pillar arrays, Line patterns) dramatically altered cell morphologies and cellular behavior.
Here, A549 human lung adenocarcinoma cells were cultured on micropillar arrays made with UV-curable polymer resin (2 micrometer diameter, 16 micrometer pitch) for 48 H and intensively investigated for the understanding of mechanotrasduction effect in (2.16) micropillar arrays. A549 cells grown on extremely stiff micropillar arrays (Young’s modulus of NOA 63 : 1.66 GPa) showed dramatic elongation effect in cell morphology. Confocal microscope image analysis reveals co-localization of actin filament withvimentin and relatively large number of nuclear deformation compared to A549 cells grown on flat NOA 63 substrate. Also, migration arrays in both in a single cell level (single cell tracking) and collective migration (wound healing assay, spreading assay) consistently confirm increased motility of cells grown in (2.16) micropillar assay. In-vivo mouse xenograft experiment also confirms increased tumor colonization in mice injected with A549 cells grown in micropillar arrays, a character acquired from mechanical stress in-vitro even persists in in-vivo environment.
We suggest that stimulation given by the (2.16) micropillar array substrate stored in cell nucleus to be passed into next generations. The relationship between the mechanical stress given by the (2.16) micropillar arrays and mechanotransduction in A549 cells will be discussed in detail.

As our understanding of the brain’s physiology and pathology progresses, increasingly sophisticated technologies are required to advance discoveries in neuroscience and develop more effective approaches to treating brain disease. There is a tremendous need for advanced materials solutions at the biotic/abiotic interface to improve the spatiotemporal resolution of neuronal recording. Organic electronic devices offer a unique approach to these challenges, due to their mixed ionic/electronic conduction, mechanical flexibility, enhanced biocompatibility, and capability for drug delivery. We designed, developed, and characterized conformable organic electronic devices in the form of transistors and electrodes to efficiently interface with the brain and acquire neurophysiological activity not previously accessible with recordings from the brain surface. These devices have facilitated large-scale rodent neurophysiology experiments and uncovered a novel
hippocampal-cortical oscillatory interaction. The biocompatibility of the devices allowed intra-operative recording from patients undergoing epilepsy surgery, highlighting the translational capacity of this class of
neural interface devices. In parallel, we are developing the high-speed electronics and embedded acquisition and storage systems required to make high channel count, chronic neurophysiological recording from animals and human subjects possible.
This multidisciplinary approach will enable the development of new devices based on organic electronics, with broad applicability to the understanding of physiologic and pathologic network activity, control of brain-machine interfaces, and therapeutic closed-loop devices.

Noninvasive transdermal delivery is a promising method with distinct advantages including patient compliance over other delivery routes. Here, hyaluronate−gold nanorod/death receptor 5 antibody (HA-AuNR/DR5 Ab) complex was developed for transdermal theranosis of skin cancer. The successful formation of the complex was corroborated by ¹H nuclear magnetic resonance, UV−vis spectroscopy, dynamic light scattering, zeta potential, and transmission electron microscopy. In vitro biological activity of the complex was verified by ELISA and MTT assay using HCT116 cancer cells. In addition, in vivo photoacoustic imaging and two-photon microscopy
clearly visualized the transdermal delivery of HA-AuNR/DR5 Ab complex through the inevitable barrier of stratum corneum in the skin. Furthermore, in vivo antitumor effect on skin cancer model mice was confirmed from statistically significant decrease of tumor-reflecting luciferase expression levels and apoptotic signals in terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay. Taken together, we could confirm the feasibility of HA-AuNR/DR5 Ab complex as a novel theranostic platform for noninvasive transdermal treatment of skin cancers.

Oral squamous cell carcinoma (OSCC) cases are major health concerns worldwide. The early detection of the disease is expected to reduce more than 50% of mortality/morbidity rate. To this end, the point-of-care devices such as easy to use biosensors may play a significant role compared to existing techniques that are costly and need trained manpower. Realization of accurate and inexpensive biosensing systems for early detection of oral cancer demands for development of robust and multifunctional material systems with excellent electron transport properties and biocompatibility. We report the synthesis of zinc oxide-reduced graphene oxide (ZnO-rGO) nanocomposite for the fabrication of electrochemical immunosensor for oral cancer biomarker ‘IL8’ detection. The immunoelectrode has been prepared by covalently immobilizing (anti-IL8) antibodies on the surface of ZnO-rGO composite thin films on ITO. The nano-immunosensor showed successful detection of IL8 at low concentration ranges i.e 100 fg mL^-1 to 5 ng mL^-1 with very high sensitivity (12.4 µA mL ng^-1), selectivity and enhanced stability. These results have been validated through in-vitro investigations using real saliva samples spiked with IL8. The obtained results indicate that ZnO-rGO composite constitutes for a potential biosensing test-bed for non-invasive onsite detection of oral cancer and can be upgraded into a device for clinical applications in future.

Current advancement in the cardiovascular field has demonstrated that transcatheter heart valve replacement is a promising alternative to traditional heart valve replacement by open heart surgery, which is expensive and invasive. Transcatheter heart valves containing flexible leaflets made from hyaluronan enhanced linear low-density polyethylene interpenetrating polymeric network (HA-LLDPE IPN) films have been shown to be hemodynamic, but the resulting IPN surfaces were not consistent. Vapor crosslinked HA-LLDPE can improve the consistency of HA surface composition. Moreover, the novel biomaterial is resistant to enzymatic degradation and is not cytotoxic. The present work describes the development of the novel HA-LLDPE fabrication process and subsequent characterization of the new materials.

The HA-LLDPE IPN formation is a multi-step process that involves swelling LLDPE film in xylenes containing silylated HA-cetrimonium complex (SHACTA) to diffuse the polysaccharide into the swollen polyethylene, vapor crosslinking the SHACTA in the LLDPE to itself above a solution containing toluene diisocyanate (TDI) and xylenes, and then reverting SHACTA back to HA. Treatment parameters were varied to study their effects on the IPN formation including SHACTA/xylenes concentration, TDI/xylenes concentration, and vapor crosslinking time and temperature. Thermal analysis helped quantify the SHACTA weight composition on treated LLDPE. Gas chromatography-mass spectrometry was implemented to determine the crosslinking vapor content. Water contact angle goniometry, infrared spectroscopy, and toluidine blue O staining were used to characterize the surface of the HA-LLDPE IPN.

Based on the collected data, TDI can effectively vapor crosslink the HA incorporated within the LLDPE, and some amount of xylenes is required during vapor crosslinking to create a durable IPN. Surface characterization reveals crosslinked HA on the LLDPE that was taken through the fabrication process, and HA was still present on LLDPE after the polymer was swollen in xylenes; this indicates that the crosslinked HA, which had been physically interlocked within the polyethylene, was not dissolved into the solvent because of the interwoven network formed between HA and LLDPE at the molecular level. In conclusion, the results demonstrate that the fabrication process involving vapor crosslinking of HA allows for the creation of an HA-LLDPE IPN. Future work will investigate the hemocompatibility potential of the biomaterial.

We are using DNA as a programmable tool for directing the self-assembly of molecules and materials. The unique specificity of DNA interactions and our ability to synthesize artificial functionalized DNA sequences makes it the ideal material for controlling self-assembly and chemical reactions of components attached to DNA sequences. In particular we are using DNA origami, large self-assembled DNA structures as a template for positioning of materials such as organic molecules, dendrimers and biomolecules.
In recent years we have developed methods for conjugation of small molecules and DNA to proteins such as antibodies. The studies are now extended to the assembly of multifunctional structures and investigation of their potential as drugs. We are furthermore using combinatins of DNA, antibodies and other proteins for development of assays for quantification of small molecule drugs in blood.

Calcium phosphate occurs naturally in bone tissue and teeth and is considered to have a good biocompatibility and a high biodegradability in biological applications.¹ Due to these properties, nanoparticles based on calcium phosphate have become a potent delivery system for a vast range of cargo molecules. This includes small molecules, DNA, RNA and proteins for in vitro and in vivo applications.^2,3 Our recent studies on therapeutic applications based on nanoparticles showed promising results in the treatment of inflammations and derived carcinogenesis, vaccination against retroviral infections, as well as viral clearance,^4-6 although the intracellular location of these nanocarriers remained uncertain yet. The aim in this project was to create a smart tool to visualize the intracellular pathway and the fate of such therapeutic nanoparticles.

The calcium phosphate nanoparticles (CaP-NPs) were synthesized according to Sokolova et al.,⁷ colloidally stabilized with fluorescence labelled poly(ethylene imine) (ATTO490LS) and loaded with a fusion protein consisting of the two fluorescent units mRFP and eGFP.⁸ The particles were characterized by dynamic light scattering (DLS) and scanning electron microscopy (SEM). HeLa cells were incubated with protein-loaded nanoparticles (2 µg of fusion-protein with 30,000 cells per well) and stained with 75 nM LysoTracker™ Deep Red for 30 minutes, washed and then kept on ice to supress the endocytic uptake until the start of the experiment. Live-cell confocal laser scanning microscopy (CLSM) was performed with a TCS SP8 system (Leica Microsystems) using a 63x/1.2 water immersion objective.

The CaP-NPs had a spherical morphology and were monodisperse with a typical diameter of 50-150 nm. The in vitro fluorescence assay confirmed a strong pH-dependency of eGFP fluorescence. At the physiological pH of 7.4, the fluorescence of both proteins is detectable. Under the conditions of an acidic pH of about 4.5-5.0 (inside a lysosome), the green fluorescence disappears due to the protonation of the chromophore of eGFP.⁹ Thus, only the red fluorescence of mRFP is detectable. Time-resolved CLSM demonstrated a successful endocytosis of protein-loaded nanoparticles into the cells, with mRFP and ATTO490LS fluorescence co-localizing with stained lysosomes. However, the fluorescence of eGFP was only detectable outside lysosomes, most likely inside of early endosomes, or at the cell membrane coming in from the outside, indicating the neutral pH at these locations. We created a smart tool to effectively visualize the intracellular pathway by using a pH-responding biosensor. This provides a versatile platform to further investigate the processing of bioactive nanocarriers inside of living cells.

REFERENCES
1. Sokolova et al., Angewandte Chemie International Edition 2008, 47, 1382-1395.
2. Kopp et al., PLoS ONE 2017, 12, e0178260.
3. Rotan et al., Beilstein Journal of Nanotechnology 2017, 8, 381-393.
4. Frede et al., Nanomedicine: Nanotechnology, Biology, and Medicine 2017, 13, 2395-2403.
5. Pastille et al., PLoS Pathogens 2017, 13, e1006649.
6. Knuschke et al., Frontiers in Immunology 2018, 9, 614.
7. Sokolova et al., Acta Biomaterialia 2013, 9, 7527-7535.
8. Klionsky et al., Autophagy 2016, 12, 1-222.
9. Kimura et al., Autophagy 2007, 3, 452-460.

ACKNOWLEDGEMENTS
The authors thank the Deutsche Forschungsgemeinschaft (DFG) for funding this project within the framework of the CRC 1093: Supramolecular Chemistry on Proteins, the ICCE for access to confocal laser scanning microscopy and Johannes Koch for technical help.

Blood, is a complex non-Newtonian fluid, comprising of three major components – erythrocytes/red blood cells (RBCs), white blood cells, and platelets suspended in a watery liquid known as the plasma. The physicochemical properties of erythrocytes such as morphology, cell volume, membrane elasticity, and protein as well as lipid compositions are regarded as the principal biomarkers pertaining to different pathophysiological states. However, the onset of certain pathological conditions such as thalassemia, sickle cell anemia, hereditary stomatocytosis etc. could significantly alter the discocytic shape of the erythrocytes and their size. The clinical procedure for the detection of these diseases requires extensive laboratorial facilities, trained manpower and time. Herein, we posit that the morphological alterations of RBCs could serve as a diagnostic marker for specific diseases; utilizing the effectiveness of the micro-scale systems.
Blood samples were collected from the peripheral veins of healthy volunteers and were centrifuged to segregate the RBCs, and upon addition of a buffer, results in the formation of a stable suspension. Poikilocytic agent (viz. Triton) was added to the diluted samples to induce deformations in the erythrocytes. Poly dimethyl siloxane (PDMS) microchannels of circular cross sections with diameters ranging from 100 – 500 µm, and length 40 mm, were fabricated using a novel, highly repeatable, soft lithography technique, with copper wires as the base mold. The diluted blood samples were then allowed to flow through the micro-channels at a controlled flow rate and the flow dynamics of the RBCs (both healthy and deformed) were monitored, and the local velocities were measured. Each experiment was repeated septuple times. It was observed that the average flow velocities, as well as the flow patterns for the healthy discoidal shaped RBCs, were significantly different from that of the deformed RBCs. We postulate that the variations could be attributed to the difference in the amount of frictional drag experienced by the healthy and the deformed erythrocytes, and affirm that this methodology could be used for the sorting of healthy red blood cells from the diseased ones.

Atomically thin membranes from 2D materials offer unique opportunities for molecular separations with high permeance and high selectivity. Here, we report the fabrication of fully functional centimeter scale nanoporous (<2-3nm) atomically thin graphene membranes for molecular separation applications. Our work focuses on systematically addressing key challenges that have limited progress in field i.e. a) scalable nanopore formation methods including in-situ approaches, b) facile support casting, c) novel defect sealing methods and d) scalable manufacturing approaches. The synthesized nanoporous atomically thin membranes show higher selectivity and permeance (up to 100 times more) than state-of-the-art polymeric membranes and can be readily used for lab-scale dialysis.

Kidambi P.R. et al. Adv. Mat. 2018
Prozorovska L. and Kidambi P.R. Adv. Mat. 2018
Kidambi P.R. et al. ACS App. Mat. & Int. 2017
Kidambi P.R. et al. Adv. Mat. 2017
Kidambi P.R. et al. Adv. Mat. 2017
Kidambi P.R. et al. Nanoscale 2017

Water-borne pathogens and their associated diseases are the largest contributors for the mortality worldwide and turn out to be a global crisis cost approximately 5500 deaths per day. These pathogens are mostly generated due to poor sanitation, industrial effluents, sedimentation of rainwater and sewage sludge. We can reduce the death rate if we are able to do rapid detection of the pathogens on-site, which would be achieved by developing a simple, user-friendly, efficient and rapid on-site-detection tool, i.e., biosensor. Here, we report a novel label-free, cost-effective paper-based E.coli biosensor by screen-printing which employs nano-architectured graphene oxide (GO) as fast electron-transfer flatland, deposited on graphene (G) screen-printed on hydrophobic paper, and thereby the bio-recognition element, lectin Concanavalin A (ConA) was immobilized on the GO to have GGO_ConA probe for bacterial sensing. Electrochemical characterization of the electrodes (G and GO) shows the fast electron-transfer with a calculated electroactive surface area of 0.16 cm². The fabricated biosensor performance was tested by Electrochemical Impedance Spectroscopy (EIS) technique. The transfer resistance (R_ct) of GGO_ConA probe increased linearly with the bacterial concentration was observed in the range of 10²-10⁸ CFU mL^-1 with an estimated limit of detection (LOD) of ~10 CFU mL^-1: This indicates the ultra-sensitivity of our biosensor platform which is 100 times more sensitive than previous studies with less time-consuming of 2-10 minutes per sample.
Our reported biosensor, being cost-effective, eco-friendly, rapid detection and ultra-sensitive for rapid detection, could be a strong candidate for point-of-care portable kits that avoids the use of expensive instruments and skilled personnel to assess water quality monitoring and diagnose the water-borne pathogens and their associated diseases.

Despite significant recent advances in materials, techniques, and implant design, orthopedic implant failure remains a significant clinical problem. In an effort to reduce implant failure rates, our group has been investigating new ways to modify the surface of implants in order to promote osseointegration, the successful integration of implant materials with normal skeletal structures. Many existing methods, such as an already FDA-approved laser rostering approach, are expensive, time inefficient, and inconsistently effective. Commercially available 316L stainless steel (316L SS) is often used for orthopedic implants due to its desirable structural properties, biocompatibility, relatively low cost, and proven success in load bearing and fixation. Recently, 3D printed 316L SS has been utilized in order to customize implants for patient-specific needs. In the work presented here, we investigated the potential of a novel, cost and time effective method of promoting osseointegration by chemically etching the implant surface with Vilella’s Reagent.

Four distinct 316L SS prototypes were investigated: (1) polished wrought 316L SS, (2) chemically etched wrought 316L SS, (3) polished 3D printed 316L SS, and (4) chemically etched 3D printed 316L SS. The 3D printed 316L SS samples were printed under identical conditions using the Renishaw AM250 manufacturing (3D printing) system. Scanning electron microscopy (SEM) was used to assess the surface roughness of each of the four prototypes. X-ray photoelectron spectroscopy (XPS) was performed on the wrought and 3D printed steel before and after etching. The contact angle of each surface was then measured using a KSV CAM 200 Contact Angle Meter. Primary dental pulp stem cells (DPSC) transduced with green fluorescent protein (GFP) were plated onto all four substrates and cultured in osteogenic medium (αMEM, 10% FBS, 0.002M Ascorbic. acid, 0.01M β Glycerol phosphate). EVOS fluorescence microscopy was used to monitor visible cell attachment daily over a period of 28 days. Confocal imaging was performed on day 28 after staining DPSC cultures with NUCLEAR-ID® Blue DNA stain and Xylenol Orange in order to assess cell proliferation and osteogenic differentiation (calcium deposition), respectively.

Consistent with our experimental postulate, SEM studies confirmed that chemical etching of both the wrought and 3D printed materials increased surface roughness as compared to the polished samples. Substrate surface chemical characterization via XPS revealed that the surface chemistry was altered following chemical etching in that there was a slight loss of Fe and Ni, and a gain of Mo. Contact angle measurements demonstrated that surface hydrophilicity was increased by the etching process on both the wrought and 3D printed samples. Fluorescent imaging demonstrated DPSC attachment to all surfaces within 48 hours of cell plating. By day 28, fluorescent imaging results showed that DPSC attachment had been enhanced on both etched over polished and 3D printed over wrought surfaces. DPSCs attached with the highest affinity to the etched 3D printed 316L SS surfaces. Confocal imaging on day 28 showed that cell proliferation and osteogenic differentiation were also maximally enhanced on the etched 3D printed 316L SS surfaces. This study demonstrates that DPSC attachment, proliferation, and osteogenic differentiation can be optimized on etched 3D printed 316L stainless steel surfaces and has clinical implications for the enhancement of orthopedic implants.

Though emerging evidence indicates that the pathogenesis of Parkinson’s disease is strongly correlated to the accumulation and transmission of α-synuclein (α-syn) aggregates in the midbrain, no anti-aggregation agents have been successful at treating the disease in the clinic. Here, we show that graphene quantum dots (GQDs) inhibit fibrillization of α-syn and interact directly with mature fibrils, triggering their disaggregation. Moreover, GQDs can rescue neuronal death and synaptic loss, reduce Lewy body and Lewy neurite formation, ameliorate mitochondrial dysfunctions, and prevent neuronto-neuron transmission of α-syn pathology provoked by α-syn preformed fibrils. We observe, in vivo, that GQDs penetrate the blood–brain barrier and protect against dopamine neuron loss induced by α-syn preformed fibrils, Lewy body/Lewy neurite pathology and behavioural deficits.

Optical upconversion that converts infrared light into visible light is of significant interest for broad applications in biomedicine. Conventional upconversion materials like rare-earth based compounds and organic dyes rely on non-linear light-matter interactions, exhibit incidence dependent efficiencies and require high power excitation. We report an infrared-to-visible upconversion strategy based on fully integrated microscale optoelectronic devices. These thin-film, ultra-miniaturized devices realize near-infrared to visible (red, yellow, green and blue) upconversion that is linearly dependent on incoherent, low-power infrared excitation. Additional features of this upconversion design include broadband absorption, wide emission spectral tunability and fast dynamics. Encapsulated, freestanding devices are transferred onto heterogeneous substrates and show desirable biocompatibilities within biological fluids and tissues. These microscale devices are implanted in behaving animals, with in vitro and in vivo experiments demonstrating their utility for optogenetic neuromodulation and photodynamic therapy. These results provide new routes for high performance upconversion materials and devices, and their unprecedented potential as optical biointerfaces.

Cancer is one of major health problems worldwide and currently the second leading cause of death in United States. Leukemia, previously the leading cause of cancer death in young people < 19 years old, has become the second cause due to dramatic advances in therapeutics and treatments. Brain cancer has now become the leading cause because of the lack of highly effective therapy. More intensive research efforts including novel strategies with clinical translation are required to bring success in the battle against this deadly disease. Glioblastoma multiforme (GBM) is the most aggressive brain tumor which has a high recurrent rate (> 90 %) and an extremely short median survival time (< 15 months). GBM local recurrence mostly within 2 cm of the original surgical lesion makes a local treatment as the most promising way to reduce GBM recurrence rates. Carmustine (bis-chloroethyl-nitrosourea - BCNU) loaded Gliadel® disc is currently the only available FDA-approved local therapy device. It is implanted along the inner walls of the brain cavity created after maximal tumor resection.¹ However, the efficacy is still not satisfactory because of a short effective period and non-conformity to the resection cavity. It releases most of BCNU drug within only 5-7 days.
Electrospinning is a highly versatile method for creating continuous fibers ranging from tens of nanometers to microns in diameter made of various composite materials, providing very high surface-to-volume ratio with a high porosity. Furthermore, using coaxial electrospinning^{2, 3}, advanced features can be obtained, such as combination of different characteristics from each polymer in different layer and encapsulating drugs in the designated layer of fibers, providing multi-functionality and efficient drug loading to the resulting membrane.
The novel approach described here uses discs formed from multi-layered core-sheath membranes (‘NanoMesh’) to provide controlled and sustained drug release for long term periods, leading to excellent in vivo results in an animal model. NanoMesh discs have been implanted into rats and their in vivo efficacy has been evaluated. Membranes with core-sheath fibers encapsulating BCNU in core were produced with different sheath thickness using coaxial electrospinning. SEM images show uniform fiber formation with average diameter of ~2.2 µm (sheath of ~ 0.36 µm), and 2.4 µm (sheath of ~ 0.56 µm). Consistent long-term release with no initial BCNU burst release was observed in vitro for up to 160 days. To evaluate in vivo efficacy, F344 rats intracranially implanted with the most malignant 9L gliosarcoma cell line were examined. All control groups (with either no treatment, no core in fibers, or no drug in fibers) rats died very early, within 12-14 days. On the other hand, rats implanted with BCNU encapsulated membrane discs remained active and healthy throughout the observation period, up to 150 days after implantation (end of trial). The survival time of treated rats increased by a factor of > 13 over the control group, which represents the largest survival improvement reported to date. Brain histology using H&E staining shows no evidence of malignant cells even at Day 150. Even better results are possible by incorporating multiple anticancer drugs into fibers, leading to synergistic effects from cell-cycle specific and non-specific drugs.
1. Lesniak, M. S.; Brem, H., Nat Rev Drug Discov 2004, 3 (6), 499-508.
2. Han, D.; Sasaki, M.; Yoshino, H.; Kofuji, S.; Sasaki, A. T.; Steckl, A. J., Journal of Drug Delivery Science and Technology 2017, 40, 45-50.
3. Han, D.; Sherman, S.; Filocamo, S.; Steckl, A. J., Acta Biomaterialia 2017, 53, 242-249.

Hollow mesoporous silica capsules (HMSC) have recently gained intense attention as drug delivery vehicles due to their biocompatibility, high loading capacity and sufficient stability in biological milieu. Herein we report on ellipsoid-shaped HMSC (aspect ratio ~ 2), synthesized using hematite particles as solid templates that were coated with a silica sol followed by acidic leaching of iron oxide. Gas sorption studies on HMSC revealed mesoscopic pores (main pore width ~38 Å) and a high surface area of 308.8 m2/g. Cell uptake and successful internalization of as-prepared HMSC was proven by confocal microscopy using human cervical cancer (HeLa) cells. The suitability of HMSC for drug delivery applications was tested by loading antibiotic (ciprofloxacin) and anticancer (curcumin) compounds as model drugs for hydrophilic and hydrophobic therapeutics. A pH dependent drug release over several days under physiological conditions at 37° C was demonstrated (UV-vis spectroscopy) in both cases, which showed the versatility of HMSC in transporting hydrophilic as well as hydrophobic drugs. Ciprofloxacin-loaded HMSC were additionally evaluated towards gram negative (E.coli) bacteria to clearly demonstrate a complete bacterial growth inhibition over 18 hours using particle concentrations of 10 µg/ml.

Vascular differentiation and formation (morphogenesis) takes place in an intricate milieu. This unique microenvironment is situated throughout the body in diverse types of healthy tissues, yet it seems to activates/inhibits similar mechanisms of the microvasculature. Two parameters of this microenvironment seem critical for blood vessel growth and stabilization: (i) the extracellular matrix, which provides critical support for vascular cell adhesion, proliferation, migration, and morphogenesis, and (ii) low oxygen concentrations (hypoxia), which is a critical factor promoting vascularization during embryonic development and tumor growth. In this talk I will present our recent efforts to develop biomaterials that mimic these physicochemical cues to understand how downstream signaling pathways impact vascular fate and assembly from progenitors and pluripotent stem cells.

Supramolecular biomaterials exploit rationally-designed non-covalent interactions to enable innovative approaches to drug formulation and delivery. For example, supramolecular interactions can be used to dynamically cross-linking polymer networks, yielding shear-thinning and self-healing hydrogels that allow for minimally invasive implantation in vivo though direct injection or catheter delivery to tissues. Herein, we discuss the preparation and application of shear-thinning, injectable hydrogels driven by non-covalent interactions between modified biopolymers (BPs) and biodegradable nanoparticles (NPs) comprised of PEG-PLA. Owing to the non-covalent interactions between PEG-PLA NPs and BPs, the hydrogels flow under applied stress and their mechanical properties recover completely within seconds when the stress is relaxed, demonstrating the shear-thinning and injectable nature of the material. The hierarchical construction of these biphasic hydrogels allows for multiple therapeutic compounds to be entrapped simultaneously and delivered with identical release profiles, regardless of their chemical make-up, over user-defined timeframes ranging from days to months. These materials enable novel approaches to immunotherapy, which rely on precise release of complex mixtures of compounds, as well as long-term treatment strategies for a variety of disease targets. Overall, this presentation will demonstrate the utility of a supramolecular approach to the design of biomaterials affording unique opportunities in the formulation and controlled release of therapeutics.

Multiphoton imaging techniques that convert low energy excitation to higher energy emission are widely used to improve signal over background, reduce scatter in subsurface imaging, and limit sample photodamage. Multiphoton imaging relies on luminescent probes able to efficiently sum the energies of 2 or more incident photons, as well as lasers powerful enough for multiphoton excitation. Lanthanide-doped upconverting nanoparticles have proven to be among the most efficient multiphoton probes, but even UCNPs with optimized lanthanide dopant levels require laser intensities that may be problematic for living systems. Here,¹ we develop protein-sized, alloyed UCNPs (aUCNPs) that can be imaged at the single particle level at laser intensities below 300 W/cm², over 300-fold lower than needed for comparably-sized doped UCNPs. Using single UCNP characterization and kinetic models of lanthanide energy transfer, we find that addition of inert epitaxial shells radically changes optimal lanthanide content from Yb³⁺, Er³⁺-doped NaYF₄ nanocrystals to fully alloyed compositions. At high levels of the emitter Er³⁺, these ions can adopt a second role to enhance the effective aUCNP absorption cross-section by desaturating sensitizer Yb³⁺ or by absorbing photons directly. Core/shell aUCNPs are brighter than comparably sized doped UCNPs at all laser intensities tested, over 4 orders of magnitude. Core/shell aUCNPs 12 nm in total diameter can be imaged with strong contrast through several millimeters of tissue in live mice using a laser intensity of just 0.1 W/cm². aUCNPs open up the possibility of using both low irradiance and low-energy excitation wavelengths for non-destructive bioimaging experiments. Additional recent work about dye-sensitized UCNP emission and UCNP-based micron-sized lasers will also be discussed.^2,3

References:
1. Low irradiance multiphoton imaging with alloyed lanthanide nanocrystals. Nature Communications 9, 3082 (2018).
2. Continuous-wave upconverting nanoparticle microlasers. Nature Nanotechnology 13, 572-577 (2018).
3. Enrichment of molecular antenna triplets amplifies upconverting nanoparticle emission. Nature Photonics 12, 402-407 (2018).

Structural materials synthesized by organisms, such as bones and shells, exhibit remarkable mechanical properties due to their hierarchical assembly of hard and soft components across the nanometer to the micron scales. While engineering analogs to these materials would open new frontiers, there is currently no route to do so that captures the key hierarchical ordering of natural composites. Here we lay the foundations for bottom-up synthesis and assembly of engineered living composites that resemble nacre using a synthetic biology approach. To do so, we engineered an artificial consortium of two Caulobacter crescentus strains. The surface-layer (S-layer) of the first ‘Displayer’ strain is engineered to permit covalent attachment of proteins, nanocrystals, and protein-based polymers to the extracellular surface. This cell surface binding is uniform, specific, and covalent, and its density can be controlled based on the location of the insertion within the S-layer. The second ‘Secreter’ C. crescentus strain is engineered to secrete a variety of protein hydrogel materials. Together the two strains program synthesis and assembly of a hydrogel enmeshing the living ‘Displayer’ cells. The properties of this living hydrogel can be changed dynamically by altering the cell-hydrogel crosslinking. Taken together, this work provides a platform for creating hierarchically-assembled living materials with switchable mechanical properties.

Since the first clinical implantation of pacemaker in 1958, the cardiac implantable medical devices have experienced a rapid evolution. While increasing population benefits from these technologies, the device-related complications such as infection, lead and battery failure are not negligible. Especially the rigid and weighty battery, not only malfunction may occur with battery depletion, but also the replacement of or recharging the batteries requires substantial surgical or technical efforts, introducing additional suffering and complexity to the patients. Therefore, the batteryless devices are on the horizon as a coming paradigm shift. To date, self-powered pacemaker and defibrillator based on triboelectric[1], piezoelectric[2] and electromagnetic-induction[3] has been proposed and demonstrated. While the feasibility of self-powered cardiac pacing has been well present, few of studies focused on the biological influence these devices caused to heart and the long-term stability of implants, which are essential to the practical applications. To address this critical issue, we present a systematic study of a cardiac energy harvester based on the ferroelectric polymer, polyvinylidene fluoride (PVDF), implanted on the left ventricle of three swine hearts for up to two months with consistent operation.
A PVDF patch with biomimetic mechanical property wire was first designed. The packaged PVDF harvester had a stable in vivo output around 1.5 V when implanted between pericardium and heart of swine. Afterwards, an up-to-two-months in vivo output of this cardiac energy harvester was recorded with device consistently operating, which exhibited a constant output from the 3 days post implantation. Meanwhile, multiple techniques, including completed blood count, cardiac enzymes test, echocardiogram, electrocardiography, as well as pressure–volume loop were used to evaluate the function of heart with implanted energy harvester. While the whole functions of heart kept normal evidenced by these techniques, no signs of toxicity or incompatibility were found from the surrounding tissues of device based on complete pathological analyses. Moreover, devices exhibited high sensitivity with stable electrical output conveying information regarding heart activity during the entire examine period and packages were also able to effectively insulate the i-NG in biological environment.
These series of in-vivo and in-vitro studies confirmed the high biological feasibility of using i-NG in vivo for cardiac energy harvesting, and thereby further enables batteryless cardiac pacing and monitoring. This research provided the first cornerstone for the future intelligent self-powered cardiac system implantation.


References:
[1] Zheng, Q., Shi, B., Fan, F., Wang, X., Yan, L., Yuan, W., Wang, S., Liu, H., Li, Z. and Wang, Z.L., 2014. In vivo powering of pacemaker by breathing-driven implanted triboelectric nanogenerator. Advanced Materials, 26(33), pp.5851-5856.
[2] Hwang, G.T., Park, H., Lee, J.H., Oh, S., Park, K.I., Byun, M., Park, H., Ahn, G., Jeong, C.K., No, K. and Kwon, H., 2014. Self-powered cardiac pacemaker enabled by flexible single crystalline PMN-PT piezoelectric energy harvester. Advanced materials, 26(28), pp.4880-4887.
[3] Zurbuchen, A., Haeberlin, A., Bereuter, L., Wagner, J., Pfenniger, A., Omari, S., Schaerer, J., Jutzi, F., Huber, C., Fuhrer, J. and Vogel, R., 2017. The Swiss approach for a heartbeat-driven lead-and batteryless pacemaker. Heart rhythm, 14(2), pp.294-299.

Water is the most significant limiting factor for crop yields. Therefore, understanding the hydrotropic response in plants (i.e., the development of roots in response to an inhomogeneous water distribution) is important to breed or design genotypes that potentially make more efficient use of a limited water supply (e.g., due to drought or to soils with low water retention). A number of studies explored the mechanism behind hydrotropism in single root tips by using osmolytes in gels and moisture in air assays , but the quantitative exploration of changes in the architecture of whole root system as a result of a known, inhomogeneous, water distribution is deceptively challenging. As far as we know, there are no technical means to generate stationary and arbitrary water distributions across a surface for the high throughput phenotyping of hydrotropic tendencies of whole root systems.

In this talk we describe a simple methodology to grow roots on a paper microfluidic device by which we control the overall amount of water available to the plant as well as where it is located with respect to the seed with a 0.83 mm spatial resolution. We use this methodology to show that, as water becomes more rare, plants of Brassica rapa concentrate a more significant fraction of their root system where water is available (up to 15% when only 1% of the available area has water, as opposed to 2% in the control). The lack of evidence for increased branching at the water sources suggest that this phenotype is purely the result of a hydrotropic, water-seeking response of the whole root system.

Root phenotypes are a great indicator for the interaction between plants and their environment, such as the uptake of water and nutrients. However, media that mimic field conditions (e.g., soil, sand) are opaque to most forms of radiation, which complicates the phenotyping of roots in vivo. On the other hand, media that are transparent (hydroponics, aeroponics, gels) do not provide field-relevant phenotypes and growing conditions. To address this issue we created “transparent soil (TS)” by a hydrogel spherification process. Differently from other “transparent soils”, our approach was uniquely designed to provide water, nutrients, and aeration to the growing roots and to allow for root phenotyping in vivo by both photography and microscopy.
The optical and mechanical properties of the gel can be readily modified, so as the effective porosity of the TS, to range between that of sand and that of loam.
We applied this TS in plant growth and evaluated its effect on root phenotypes and stress response on plants such as Glycine max, Brassica rapa and Arabidopsis thaliana. Compared to hydroponic media, this TS provides root phenotypes in Glycine max that are significantly more similar to those observed in field soil and avoid stress
response from root hypoxia.
Lastly we show how the tailorability of this new materials platform allows for the investigation of how root development is affected by soil heterogeneities, e.g, gradients in mechanical properties (“thigmotropism”) or water availability (“hydrotropism”). We expect that this new application of hydrogels will provide, through synthetic polymers and their functionalization, broad possibilities for the study of GxE interactions in root development, as well as the modeling of the rhizosphere.

Bone as a high-performance functional material encounters various mechanical forces such as tensile, shear and periodic i.e. fatigue stresses which make it prone to the mechanical damages specially microcracks. the duly diagnosis of bone microcracks is crucial to estimate the fracture and fragility risk factors and to conduct the proper intervention. Several clinically used imaging modalities are currently being investigated for non- invasive three-dimensional assessment of bone microdamage in vivo. Among them, computed tomography (CT) imaging is an excellent candidate due to being clinically amenable and its capability to provide information on the anatomical features rapidly. However, the confounding radiographic attenuation from bone instrument make the bone damage detection difficult while the low spatial resolution of the machine does not allow detection of damage on the micrometer scale.
Nanoparticles composed of high atomic number materials can partially resolve the issues associated with strong x-ray attenuation from calcium in the conventional CT imaging machines. When combined with advances in the CT instrumentation, the best outcome is expected. Specifically, the incorporation of photon counting detectors resulted in the multispectral CT technology which enables the detection of exogenous contrast materials with minimal background from hard tissues. Depending on the various inherent K-edge of materials, they can be discriminated when applying fast voltage switching of the X-ray source. Materials whose K-edge absorption band falls in the range of peak tube potential of 80-140 kVp are especially suitable since they can bypass beam hardening effect in these practical beam energy ranges. It was indicated that the elements with Z=64-73 have the potential to enhance the x-ray contrast effectively as compared to iodine while higher Z elements would reduce the photon counts sensed by the detectors (such as Au, Bi, etc).
Hf with Z=72 is an emerging radiopaque material with well-positioned K-edge energy (65.3 keV) has been rarely investigated for contrast agent in CT imaging especially K-edge imaging. Herein, we disclose for the first time the synthesis of the ligand-modified sub 5 nm HfO₂ nanoparticles for the sensitive detection of bone microdamage. Furthermore, these nanoparticles were targeted to bone minerals by the introduction of an aminopolycarboxylic chelating agent. At the site of the microcracks, a fracture in hydroxyapatite, the major bone constituent, would result in the exposure of freshly charged ions due to the breakage of atomic bonds in the matrix. Hence, the nanoparticles targeted to the more concentrated minerals would ‘light up’ the microcrack site. We demonstrated that with the distinct spectral properties, HfO₂ nanoparticles would show in ‘color’ the location and anatomy of the microcrack. Moreover, the sub 5 nm size of these nanoparticles makes it feasible for them to travel through the nutrient transport canals (e.g. Haversian canals, canaliculi, Volkmann’s canals, etc.) in the muscle near the bone and microcrack itself. We indicated in the in vivo studies that these nanoparticles could effectively indicate the microcrack anatomy in the rat’s tibia in the spectral CT while the non-targeted control nanoparticles non- specifically accumulated in the muscle. Furthermore, intramuscular injection of the nanoparticles did not induce any adverse effects on the liver enzyme functions and more importantly histopathologically. Therefore, these ligand-directed HfO₂ nanoparticles would open new avenues for the safe, non-destructive, sensitive bone microdamage detection in vivo with potential clinical translation.

Separation and other forms of microfluidic manipulation of white blood cells for immunology and related applications is critical for more specific diagnostics and future treatments. We report on the studies of mechanical and electrical properties of bio-hybrid of white blood cells (WBCs) intermixed with gold nanoparticles (AuNPs) through dielectrophoresis (DEP) measurements. Studies of WBCs are performed with both, standard form of WBCs, and in gold nanoparticles (AuNPs) enriched form. DEP devices are microfabricated in the form of metal-on-glass microelectrode arrays, with the shape of the elements of DEP devices numerically modelled following broadly accepted elementary physical properties of white blood cells. Specifically, indium tin oxide (ITO) was evaporated onto glass to 150 nm thickness, then lithographically patterned and cured at 400 deg C to modify its transparency and achieve higher conductivity. Afterwards, individual DEP chips are used for the deposition of small volumes of cell-AuNP culture suspension into the DEP electrode region. Electrodes are connected to a function generator and wired to the source of an AC current, where electric field and frequency could both be controlled. Jurkat and THP-1 cell types are tested so far. We have followed hypothesis that mechanical modulus value and its possible changes are related to the cells' activation, which is considered as relevant in some elements of body's immune response. Additional motivation for this set of experiments is the link between cancer and WBC’s deformability and other mechanical properties. Two main types of studies are reported here. In the first, frequency sweeps are performed at a fixed voltage (10V), and the crossover frequency is measured to be 18.0+/- 2.00 kHz for THP-1 cells and 77.5 +/- 4.50 kHz for Jurkat cells. In the second series of studies, elastic modulus is determined following measurements of aspect and stretch ratios, with THP-1 modulus in the range of 430 Pa, and Jurkat cells modulus at around 530 Pa. The introduction of Au NPs allows for the modification of the elastic modulus values by a factor varying from 1.5 to 4, depending on the reagents used when Au NPs are added (PEG with varied concentration, citrate, others), and depending on the concentration of Au NPs. Measurements so far indicate that at least a portion of Au NPs are absorbed into WBCs, while the rest are either close to WBC outer membrane or “free floating” in the cell culture suspension. As a potentially encouraging news, the addition of controllable amount of AuNPs doesn't negatively impact the overall deformability of WBCs, despite the modifications to the elastic modulus values. This hybrid, bio-materials centered system of white blood cells and gold nanoparticles is a subject of continued studies, and will likely serve as a precursor for better understanding of related systems of interest in biomaterials and immunology.

Cancer, the major public health issue, has become the leading cause of death in China since 2010. Traditional chemotherapy is still a leading approach in current cancer treatment to suppress tumor proliferation and prolong patient survival by eliminating or inhibiting cancer cells through anticancer drugs. Nanomedicine has become rapidly growing area of medical research for cancer treatment, as they can efficiently carry and deliver therapeutic agents, imaging probes or biological materials to the targeted tumor sites by enhanced permeability and retention (EPR) effect. However, the safety of nanoparticles is essential for clinical application. Polyphenols, compounds found in our diet, could help to prevent degenerative diseases such as cancer and cardiovascular diseases. Therefore, polyphenols based nanomaterials can be employed as excellent candidates for nanomedicine. Moreover, the polyphenols would seem to have great potential to promote the efficacy of anticancer drugs. We constructed a self-assembled nanoplatform with the help of interactions between polyphenols and Fe³⁺ ion. Epigallocatechin gallate (EGCG) and PEG polyphenols were employed as the polyphenols, and anticancer drug doxorubicin (DOX) was encapsulated in the core of the nanoparticles for cancer treatment. The size of nanoparticles could be controlled from 30 nm to 150 nm by tuning the Fe³⁺ ion concentration. More interestingly, the nanoplatform can be used for in vivo fluorescence imaging with NIR dye modification and T₁ magnetic resonance imaging (Fe³⁺), which is useful for cancer diagnosis. The nanoparticles are able to promote tumor treatment efficacy and prolong mice survival time compared with free drug alone. This theranostics nanoplatform establishes a novel promising approach to design next generation nanoparticles for cancer treatment.